The outbreak of novel coronavirus (COVID-19) infections in 2019 is in dire need of finding potential therapeutic agents. In this study, we used molecular docking to repurpose HIV protease inhibitors and nucleoside analogues for COVID-19, with evaluations based on docking scores calculated by AutoDock Vina and RosettaCommons. Our results suggest that Indinavir and Remdesivir possess the best docking scores, and comparison of the docking sites of the two drugs reveal a near perfect dock in the overlapping region of the protein pockets. After further investigation of the functional regions inferred from the proteins of SARS coronavirus, we discovered that Indinavir does not dock on any active sites of the protease, which may give rise to concern in regards to the efficacy of Indinavir. On the other hand, the docking site of Remdesivir is not compatible with any known functional regions, including template binding motifs, polymerization motifs and nucleoside triphosphate (NTP) binding motifs. However, when we tested the active form (CHEMBL2016761) of Remdesivir, the docking site revealed a perfect dock in the overlapping region of the NTP binding motif. This result suggests that Remdesivir could be a potential therapeutic agent. Clinical trials still must be done in order to confirm the curative effect of these drugs.

Minimal plasmids play an essential role in many intermediate steps in molecular biology. They can for example be used to assemble building blocks in synthetic biology or be used as intermediate cloning plasmids that are ideal for PCR-based mutagenesis methods. A small backbone also opens up for additional unique restriction enzyme cloning sites. Here we describe the generation of pICOz, a 1185 bp fully functional high-copy cloning plasmid with an extended multiple cloning site (MCS). To our knowledge, this is the smallest high-copy cloning vector ever described.

Background: Studies employing next-generation sequencing (NGS) show that the oral fungal community (mycobiome) is far more complex than hitherto thought. However, the role of the oral mycobiome in health and disease, including oral carcinogenesis, has not been explored. Objective: To characterize the mycobiome associated with oral squamous cell carcinoma (OSCC). Methods: Tissue biopsies [cases: 25 OSCC; controls: 27 intra-oral fibro-epithelial polyp (FEP)] were collected from oral and maxillofacial units in Sri Lanka. Total DNA was extracted and subjected to sequencing of the fungal ITS2 region using Illumina’s 2x300 bp chemistry. High quality, non-chimeric merged reads were classified to species level using a BLASTN-algorithm with UNITE’s named species sequences as reference. Downstream analyses were performed using QIIME and LEfSe. Results: 364 species representing 160 genera and 2 phyla (Ascomycota and Basidiomycota) were identified, with Candida and Malassezia making up 48% and 11% of the average mycobiome, respectively. However, only 5 species and 4 genera were detected in ≥50% of the samples. The species richness and diversity were significantly lower in OSCC. At the genus level, Candida, Hannaella and Gibberella were overrepresented in OSCC while Alternaria and Trametes were more abundant in FEP. Species-wise, C. albicans, C. etchellsii and Hannaella luteola-like species were enriched in OSCC while Malassezia restricta, Aspergillus tamarii, Alternaria alternate, Cladosporium halotolerans, and Hanseniaspora uvarum-like species were the most significantly abundant in FEP. Conclusions: A dysbiotic mycobiome dominated by C. albicans was found in association with OSCC. Whether this dysbiosis plays a role in oral carcinogenesis warrants further investigation.

Current theories of autoimmunity are diverse, sometimes contradictory, and suffer from incompleteness. Although substantial evidence exists that adaptive and innate immunity, sex, genetic predisposition, and the microbiome all play essential roles in autoimmune disease etiologies and pathogenesis, and that antigen processing is altered during disease induction, no existing theory integrates all of these factors through a single, coherent mechanism. In an attempt to focus the field on the need to elucidate such an integrative mechanism, I propose one possibility here that, if nothing else, helps to identify the nature of the problems that need to be addressed. My theory is that autoimmune diseases are induced by normal immunological responses to unique pairs of complementary antigens, at least one of which is a molecular mimic of a host target. Each antigen in the complementary pair induces a complementary immune response (T or B cell); although each immune response is idiotypic in origin, the antigenic complementarity results in what appears to be an idiotype-anti-idiotype relationship between the responses. Additionally, because of the antigenic complementarity, each immune response mimics one of antigens, abrogating the distinction between self and non-self. If at least one of the antigens mimics a host antigen, then the resulting immunological civil war spreads to a host tissue. Complementary antigens also alter antigen processing so that antigens that would normally be proteolytically digested are presented by the major histocompatibility complex (MHC) to T and B cell receptors inducing a cross-reactive immune response. The resulting civil war is supported by the innate immune system due to the complementarity of the initiating antigens.. Complementary antigens stimulate synergistic toll-like receptors (TLR) and/or nucleotide-binding oligomerization receptors (NOD) resulting in up-regulation of cytokine production and further stimulation of the adaptive immune response. Because the immune responses (e.g., antibodies) mimic the initiating antigens, this synergistic activation of innate immunity becomes chronic. Additionally, TLR and NOD function are highly sensitive to sex hormones, some becoming up-regulated and some down-regulated in the presence of either testosterone or estrogens. This sensitivity explains how sex modifies susceptibility to autoimmune diseases. Genetic mutations in TLR, NOD and MHC further alter antigen presentation and the degree to which antigens stimulate an immune response explaining how genetics also modifies susceptibility. Finally, sex hormones also alter the host microbiome, which in turn modulates autoimmune disease risk by shaping the immunological nature of self and by mediating susceptibility to microbial infection. Moreover, it appears that the microbiome camouflages itself from the immune system by mimicking the host antigenic repertoire; the mimicry between the antigens of the microbiome and the host results in selective attacks on microbiome constituents concomitant with any autoimmune attack on host tissues. This antigenic complementarity theory thereby integrates all major elements known to affect, or be affected by, autoimmune diseases and provides a set of testable implications.

Echinococcus granulosus is a harmful cestode parasite which could cause Cystic Echinococcosis in humans, various livestock species and wild animals. Calmodulin (CaM), a Ca²⁺ sensor protein, is widely expressed in eukaryotes and mediates a variety of cellular signaling activities. In our study, the CaM in Echinococcus granulosus (rEgCaM) was successfully cloned and the molecular and biochemical characterizations of rEgCaM were identified. The results showed that rEgCaM was a highly conserved calcium-binding protein, consisting of 149 amino acids. Immunoblot analysis revealed that rEgCaM could be identified using E. granulosus infected sheep serum. The usage of rEgCaM as antigen was evaluated by indirect ELISA which exhibited a high sensitivity of 90.3% but low specificity (47.1%). rEgCaM was mainly located in the tegument tissues and parenchymal region of protoscoleces, the tegument and inner body of adult worm and predominantly expressed in the germinal layer. The mRNA expression of rEgCaM in PSCs were gradually decreased with the increase death of PSCs. In electrophoretic mobility tests and ANS assays, rEgCaM showed a typical calcium-binding protein characteristics. This is the first report on CaM from E. granulosus and rEgCaM is considered to be involved in some important biological function of E. granulosus as a calcium-binding protein.

Studies on the possible association between bacteria and oral squamous cell carcinoma (OSCC) remain inconclusive, largely due to methodological variations/limitations. The objective of this study was to characterize the species composition as well as functional attributes of the bacteriome associated with OSCC. DNA obtained from 20 fresh OSCC biopsies (cases) and 20 deep-epithelium swabs (matched control subjects) were sequenced for the V1-V3 region using Illumina’s 2x300 bp chemistry. High quality, non-chimeric merged reads were classified to species level using a prioritized BLASTN-algorithm. Downstream analyses were performed using QIIME, PICRUSt, and LEfSe. Fusobacterium nucleatum subsp. polymorphum was the most significantly overrepresented species in the tumors followed by Pseudomonas aeruginosa and Campylobacter sp. Oral taxon 44, while Streptococcus mitis, Rothia mucilaginosa and Haemophilus parainfluenzae were the most significantly abundant in the controls. Functionally, genes involved in bacterial mobility, flagellar assembly, bacterial chemotaxis and LPS synthesis were enriched in the tumors while those responsible for DNA repair and combination, purine metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, ribosome biogenesis and glycolysis/gluconeogenesis were significantly associated with the controls. This is the first epidemiological evidence for association of F. nucleatum and P. aeruginosa with OSCC. Functionally, an “inflammatory bacteriome” is enriched in OSSC.

At nuclear scale, we present three heuristic relations pertaining to strong and electroweak coupling constants. With these relations, close to beta stability line, it is possible to study nuclear binding energy with a single energy coefficient of magnitude $\left(\frac{1}{{\alpha }_s}\right)\left[\frac{e^2}{4\pi {\epsilon }_0{R}_0}\right]\approx 10.0\text{MeV}\text{.}$ With reference to up and down quark masses, it is also possible to interpret that, nuclear binding energy is proportional to the mean mass of $\left[\left(2m_u+m_d\right)\text{and}\left(m_u+2m_d\right)\right]\approx 10.0\text{MeV}.$

A new type of low cost, environmentally friendly wood-plastic composites (WPC) containing carbon nanotubes(CNT)of low content 0%, 0.05wt%, 0.1wt% and 0.15wt%, wood fibers of 14wt% and polymer PES of 86wt% was manufactured by the selective laser sintering (SLS) approach of 3D printing. The experimental results showed that the incorporating of CNTs could obviously increase the mechanical properties of the wood/PES composites material. The tensile strength, bending strength and elasticity modulus were 76.3%, 227.9% and 128.7% higher with 0.1wt% CNTs than without CNTs. And the mechanical properties of specimens firstly increased and then decreased with the increasing contents of CNTs. The SEM results of the specimens’ fracture morphology indicated that the preferable bonding interfaces between wood flour grains and PES grains were achieved by adding CNTs to the composites. There are two reasons to explain why the composites possessed the superior mechanical properties: CNTs could facilitate the laser sintering process of wood plastic composites due to their thermal conductivities; also, CNTs could directly reinforce the WPC composites as reinforcement.

Adeno-associated viruses (AAVs, genus dependoparvovirus) are promising gene therapy vectors. In strains AAV1-12, the capsid gene VP1 encodes a recently discovered protein, MAAP, in an overlapping frame. MAAP binds the cell membrane by an unknown mechanism. We discovered that MAAP is also encoded in bovine AAV and in porcine AAVs (which have shown promise for gene transfer into muscle tissues), in which it is probably translated from a non-canonical start codon. MAAP is predicted to be mostly disordered except for a predicted C-terminal, membrane-binding amphipathic α-helix. MAAP has a highly unusual composition. In particular, it lacks internal methionines, and is devoid of tyrosines in most strains. Unexpectedly, we discovered that the N-terminus of VP1 also lacks several amino acids. In all AAVs that encode MAAP, the first 200 aas of VP1 are devoid of internal methionines, probably owing to a selection against ATG codons that could prevent translation of MAAP and of capsid isoforms (VP2, VP3). The N-terminus of VP1 also lacks cysteines, likely to avoid the formation of disulfide bridges when it becomes exposed outside of the capsid during post-endocytic trafficking. Finally, the region common to VP1 and VP2 lacks tyrosine in the vast majority of AAVs that encode MAAP. Avoiding these "forbidden" aas in MAAP and VP1 when creating recombinant AAV capsids might increase the efficiency of capsid design. Conversely, the presence of "forbidden" aas in some rare strains probably indicates that they have unusual properties that could help us understand the viral cycle.

An in vitro medium-throughput screen using M. tuberculosis H37Rv was employed to screen an in-house library of structurally diverse compounds for antimycobacterial activity. From this initial screen, eleven 7-substituted coumarin derivatives with confirmed monoamine oxidase-B and cholinesterase inhibitory activities, demonstrated growth inhibition of more than 50% at a 50 µM concentration. This prompted further exploration of all the 7-substituted coumarins in our library, nineteen in total, as potential antimycobacterial agents. Four derivatives showed promising antimycobacterial activity with MIC₉₉ values of 8.31 – 29.70 µM and 44.15 – 57.17 µM on M. tuberculosis H37Rv in independent assays using Gaste-Fe and 7H9 + OADC media, respectively. These compounds were found to bind to albumin which may explain the variations in MIC between the two assays. Preliminary antimycobacterial evaluation of moxifloxacin resistant M. tuberculosis show that these compounds are able to maintain their activity in fluoroquinolone resistant mycobacteria. Analysis of structure activity relationships for antimycobacterial versus neuronal enzyme inhibitory activity indicate that structural modification on position 4 and/or 7 of the coumarin scaffold may be utilized to improve selectivity towards either inhibition of neuronal enzymes or antimycobacterial effect. Cytotoxicity evaluations of the compounds indicate moderate cytotoxicity with slight selectivity towards mycobacteria. Further neuroprotective assays on SH-SY5Y human neuroblastoma cells indicate significant neuroprotection for selected compounds irrespective of their neuronal enzyme inhibitory properties. These coumarin molecules are thus interesting lead compounds that may provide insight into the design of new antimicrobacterial and/or neuroprotective agents.

A SARS-CoV receptor-binding domain (RBD) recombinant protein was developed and manufactured under current good manufacturing practices in 2016. The protein known as RBD219-N1 when formulated on Alhydrogel®, induced high-level neutralizing antibodies and protective immunity with minimal immunopathology in mice after a homologous virus challenge with SARS-CoV (MA15 strain). In this report, we examined published evidence in support of whether the SARS-CoV RBD219-N1 could be repurposed as a heterologous vaccine against Coronavirus Infectious Disease (COVID)-19. Our findings include evidence that convalescent serum from SARS-CoV patients can neutralize SARS-CoV-2. Additionally, a review of published studies using monoclonal antibodies (mAbs) raised against SARS-CoV RBD and that neutralize the SARS-CoV virus in vitro, finds that some of these mAbs bind to the receptor-binding motif (RBM) within the RBD, while others bind to domains outside this region within RBD. This information is relevant and supports the possibility of developing a heterologous SARS-CoV RBD vaccine against COVID-19, especially due to the finding that the overall high amino acid similarity (82%) between SARS-CoV and SARS-CoV-2 spike and RBD domains is not reflected in RBM region (59%). However, the high similarity (94%) in the region outside of RBM offers the potential of conserved neutralizing epitopes between both viruses.

Carbon supported nanoparticles of monometallic Ni catalyst and binary Ni-Transition Metal (Ni-TM/C) electrocatalytic composites were synthesized via chemical reduction method, where TM stands for the doping elements Fe, Co, and Cu. The chemical composition, structure and morphology of the Ni-TM/C materials were characterized by X-ray photoelectron spectroscopy (XPS), X-ray diffractometry (XRD), transmission electron microscopy (TEM), scanning transmission electron microscopy (STEM) and energy-dispersive X-ray spectroscopy (EDS). The electrochemical properties towards hydrogen oxidation reaction in alkaline medium were studied using the rotating disc electrode and cycling voltammetry methods. A significant role of the TM dopant in the promotion of the hydrogen electrooxidation kinetics of the binary Ni-TM/C materials were revealed. A record-high in exchange current density value of 0.060 mA cm²_Ni was measured for Ni₃Fe₁/C, whereas the monometallic Ni/C counterpart has only shown 0.039 mA cm²_Ni. In order to predict the feasibility of the electrocatalysts for hydrogen chemisorption, density functional theory was applied to calculate the hydrogen binding energy and hydroxide binding energy values for bare Ni and Ni₃TM₁.

Zinc finger proteins (ZFP) play important roles in cellular processes. The DNA binding region of ZFP consists of 3 zinc finger DNA binding domains connected by amino acid linkers, the sequence TGQKP connects ZF1 and ZF2, and TGEKP connects ZF2 with ZF3. Linkers act to tune the zinc finger protein in the right position to bind its DNA target, the type of amino acid residues and length of linkers reflect on ZF1-ZF2-ZF3 interactions and contribute to the search and recognition process of ZF protein to its DNA target. Linker mutations and the affinity of the resulting mutants to specific and nonspecific DNA targets were studied by MD simulations and MM_GB(PB)SA. The affinity of mutants to DNA varied with type and position of amino acid residue. Mutation of K in TGQKP resulted in loss in affinity due to the loss of positive K interaction with phosphates, mutation of G showed loss in affinity to DNA, WT protein and all linker mutants showed loss in affinity to a nonspecific DNA target, this finding confirms previous reports which interpreted this loss in affinity as due to ZF1 having an anchoring role, and ZF3 playing an explorer role in the binding mechanism. The change in ZFP-DNA affinity with linker mutations is discussed in view of protein structure and role of linker residues in binding.

Many studies discuss carbon-based materials because of the versatility of its element. They include different opinions for scientific problems and discuss fairly at convincing and compelling levels within the scope and application. A gas-state carbon atom converts into various states depending on its conditions of processing. The electron transfer mechanism in the gas-state carbon atom is responsible to convert it into various states, namely, graphite, nanotube, fullerene, diamond, lonsdaleite and graphene. The shape of ‘energy trajectory’ enables transferring electrons from the left- and right-sides of an atom is like a parabola. That ‘energy trajectory’ is linked to states (filled state and suitable nearby unfilled state) where force-exertion along the poles of transferring electrons is remained balance. So, the mechanism of originating different states of a gas-state carbon atom is under the involvement of energy first. This is not the case for atoms executing confined inter-state electron-dynamics as the force is involved first. Graphite-, nanotube- and fullerene-state atoms ‘partially evolve partially develop’ (form) their structures. These possess one-dimensional, two-dimensional and four-dimensional ordering of atoms, respectively. Their structural formation also comprises ‘energy curve’ having a shape-like parabola. Transferring suitable filled state electron to suitable nearby unfilled state is under a balance force exerting along the poles. The graphite structure under only attained-dynamics of atoms can also be formed but in two-dimension. Here, binding energy between graphite-state carbon atoms is for a small difference of exerting forces along their opposite poles. Structural formation in diamond, lonsdaleite and graphene atoms involve energy to gain required infinitesimal displacements of electrons through which they maintain orientationally-controlled exerting forces along dedicated poles. In this study, the growth of diamond is found to be south to east-west (ground) where atoms bound ground to south. Thus, diamond atoms merge for a tetra-electron ground to south topological structure. Lonsdaleite atoms merge for a bi-electron ground to just-south topological structure. The growth of graphene is found to be north to ground where atoms bound ground to north. Thus, graphene atoms merge for a tetra-electron ground to north topological structure. Glassy carbon exhibits layered-topological structure where, tri-layers of gas-, graphite- and lonsdaleite-state atoms successively bind in repetitive order. Nanoscale hardness is also sketched based on different force-energy behaviors of different state carbon atoms. Here, structure evolution in each carbon state atom explores its own science.

The transnational spread of coronavirus (2019-nCoV) first detected in Wuhan is causing global panic; thus, accelerated research into clinical intervention is of high necessity. The spike glycoprotein structure has been resolved, and its affinity to human angiotensin-converting enzyme 2 (ACE-2) has been experimentally validated. Here, using computational methods, a metastable conformation of 2019-nCoV-RBD/ACE-2 complex has been revealed and FDA-database of approved drugs have been docked into the interface. Darunavir has been discovered as high ligand affinity candidate capable of disrupting communication between 2019-nCoV-RBD and ACE-2. Darunavir, in addition to its previously known anti-HIV protease inhibitor is now repurposeable for the treatment 2019-nCoV disease acting via disruption of cellular recognition, binding and invasion.

There is much interest in effective copper chelators to correct copper dyshomeostasis in neurodegenerative and oncological diseases. In this study, a recombinant fusion protein for expression in E. coli cells was constructed from glutathione-S-transferase (GST) and the N-terminal domain (ectodomain) of human high affinity copper transporter CTR1 (hNdCTR1), which has three metal-bound motifs. Several biological properties of the GST-hNdCTR1 fusion protein were assessed. It was demonstrated that in cells, the protein was prone to oligomerization, formed inclusion bodies and displayed no toxicity. Treatment of E. coli cells with copper and silver ions reduced cell viability in a dose- and time-dependent manner. Cells expressing GST-hNdCTR1 protein demonstrated resistance to the metal treatments. These cells accumulated silver ions and formed nanoparticles that contained AgCl and Ag0. In this bacterial population, filamentous bacteria with length about 10 μm were often observed. The possibility for the fusion protein carrying extracellular metal binding motifs to integrate into the cell’s copper metabolism and its chelating properties are discussed.

Weed competitive ability (WCA) is a desirable key trait for the improvement of grain yield under direct-seeded rice (DSR) and the aerobic rice ecosystem. The present study targeted screening of 167 introgression lines (ILs) of a Green Super Rice (GSR) IR2-6 population derived from a cross between Weed Tolerant Rice 1 (WTR1) as the recipient parent and Y134 as the donor parent developed at IRRI for weed competitiveness in screen house conditions (SHC). The ILs were phenotyped for WCA traits such as early seed germination (ESG) and early seedling vigor (ESV) in Petri dishes and pot experiment conditions. The results of phenotypic variance revealed ESG-related traits, especially first germination count (1st GC) that positively correlated with second germination count (2nd GC), germination percentage (GP), total dry weight (TDW), total fresh weight (TFW), and vigor index (VI-1), whereas, in ESV, all the traits were positively correlated with each other except for three traits: root dry weight (RDW), 1st GC, and GP-2. The ESG and ESV traits are vital for weed competitiveness. A 6K SNP array was used to study the genetic association for the WCA traits. Forty-four QTLs for WCA traits were mapped on all chromosomes (except on chromosomes 4 and 8) through single marker analysis (SMA). Out of 44 QTLs, 29 were associated with ESG traits and 15 with ESV traits, with LOD scores of 2.93 to 8.03 and 2.93 to 5.04 and explained phenotypic variance ranging from 7.85% to 19.9% and from 7.85% to 13.2%, respectively. However, 31 QTLs were contributed by a negative additive allele from Y134, whereas a positive additive allele was contributed by WTR1 in 13 QTLs. Among them, two QTL hotspot regions were mapped on chromosome 11 (24.7-27.9 Mb) and chromosome 12 (14.8-17.4 Mb). The majority of the QTLs related to WCA traits were grouped into two QTL hotspots: QTL hotspot-I (qAFW11.1, qFC11.1, qFC11.2, qSC11.1, qGP-111.1, qGP-111.2, qTFGS11.1, qVI-111.1, and qVI-111.2) and QTL hotspot-II (qFC12.1, qFC12.2, qSC12.1, qFC12.2, qGP-112.1, qGP-112.2, qTFGS12.1, qTFGS12.2, qVI-112.1, qIV12.2, qFC12.1, and qGC12.2), and a few of them were co-localized on chromosomes 11 and 12. Further, we fine-tuned in the genomic regions of QTL hotspots and identified a total of 13 putative candidate genes on chromosomes 11 and 12 collectively. The present study is the first report on the genetic basis of WCA-related traits and the co-localized QTLs, which could be highly valuable in future breeding programs aiming to improve WCA in rice.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), a novel evolutionarily divergent RNA virus etiological agent of COVID-19, is responsible for present devastating pandemic respiratory illness. To explore the genomic signatures, we comprehensively analyzed 2,492 complete and/or near-complete genome sequences of SARS-CoV-2 strains reported from across the globe to the GISAID database up to 30 March 2020. Genome-wide annotations revealed 1,407 nucleotide-level mutations at different positions throughout the entire genome of SARS-CoV-2. Moreover, nucleotide deletion analysis found nine deletions throughout the genome, including in polyprotein (n=6), ORF10 (n=1) and 3´-UTR (n=2). Evidence from the systematic gene-level mutational and protein profile analyses revealed a large number of amino acid (aa) substitutions (n=722), making the viral proteins heterogeneous. Notably, residues of receptor-binding domain (RBD) having crucial interactions with angiotensin-converting enzyme 2 (ACE2), and cross-reacting neutralizing antibody were found to be conserved among the analyzed SARS-CoV-2 strains, except for replacement of Lysine with Arginine at 378 position of the cryptic epitope of a Shanghai isolate, hCoV-19/Shanghai/SH0007/2020 (EPI_ISL_416320). Our method of genome annotation is a promising tool for monitoring and tracking the epidemic, the associated genetic variants, and their implications for the development of effective control and prophylaxis strategy.

In this work α-synuclein amyloid fibrils, formation of which is a biomarker of the Parkinson’s disease, were investigated with the use of fluorescent probe thioflavin T (ThT). Experimental conditions of the protein fibrillogenesis were chosen so that a sufficient number of continuous measurements can be performed to characterize and analyze all stages of this process. The reproducibility of fibrillogenesis and the structure of the obtained aggregates (that is a critical point for their further investigation) were proved using a wide range of physical-chemical methods. For determination of ThT—α-synuclein amyloid fibrils binding parameters sample and reference solutions were prepared with the use of equilibrium microdialysis. By absorption spectroscopy of these solutions ThT—fibrils binding mode with the binding constant about 10⁴ M⁻¹ and stoichiometry of ThT per protein molecule about 1:8 was observed. Fluorescence spectroscopy of the same solutions with the subsequent correction of the recorded fluorescence intensity on the primary inner filter effect allowed to determine another mode of ThT binding to fibrils with the binding constant about 10⁶ M⁻¹ and stoichiometry about 1:2500. Analysis of photophysical characteristics of the dye molecules bound to the sites of different binding modes allowed to assume the possible localization of these sites. Obtained differences in the ThT binding parameters to amyloid fibrils formed from α-synuclein and other amyloidogenic proteins, as well as in the photophysical characteristics of the bound dye, confirmed the hypothesis of amyloid fibrils polymorphism.

We analyzed the affinity-proteomics data of saliva absorbed to plate-bound Spike protein of SARS-CoV-2, and identified major virus-binding proteins as MUC7, MUC5B, DMBT1, and neutrophil defensins. Furthermore, we found that saliva from healthy donors inhibited the binding of Spike-protein-specific polyclonal antibodies to Spike antigen. These data suggest that the Spike protein’s glycoprotein-binding domains (GBD) may be targeted to block virus adherence or entry of SARS-CoV-2.

Bacteria currently included in Rhizobium leguminosarum are too diverse to be considered a single species, so we can refer to this as a species complex (the Rlc). We have found 429 publicly available genome sequences that fall within the Rlc and these show that the Rlc is a distinct entity, well separated from other species in the genus. Its sister taxon is R. anhuiense. We constructed a phylogeny based on concatenated sequences of 120 universal (core) genes, and calculated pairwise average nucleotide identity (ANI) between all genomes. From these analyses, we concluded that the Rlc includes 18 distinct genospecies, plus 7 unique strains that are not placed in these genospecies. Each genospecies is separated by a distinct gap in ANI values, usually at around 96% ANI, implying that it is a 'natural' unit. Five of the genospecies include the type strains of named species: R. laguerreae, R. sophorae, R. ruizarguesonis, "R. indicum" and R. leguminosarum itself. The 16S ribosomal RNA sequence is remarkably diverse within the Rlc, but does not distinguish the genospecies. Partial sequences of housekeeping genes, which have frequently been used to characterise isolate collections, can mostly be assigned unambiguously to a genospecies, but alleles within a genospecies do not always form a clade, so single genes are not a reliable guide to the true phylogeny of the strains. We conclude that access to a large number of genome sequences is a powerful tool for characterising the diversity of bacteria, and that taxonomic conclusions should be based on all available genome sequences, not just those of type strains.

Several clinically useful anticancer drugs selectively kill cancer cells by inducing DNA damage; the genomic instability and DNA repair defects of cancer cells make them more vulnerable than normal cells to the cytotoxicity of DNA-damaging agents. Because epoxide-containing compounds can induce DNA damage, we have used the MTT assay to evaluate the selective cytotoxicity of three epoxyalkyl galactopyranosides against A549 lung cancer cells and MRC-5 lung normal cells. Compound (2S,3S)-2,3-Epoxydecyl 4,6-O-(S)-benzylidene-β-D-galactopyranoside (EDBGP) showed the highest selective anticancer activity and was selected for mechanistic studies. After observing that EDBGP induced cellular DNA damage (comet assay), we found that cells deficient in nucleotide excision repair were hypersensitive to the cytotoxicity of this compound; this suggests that EDBGP may induce bulky DNA adducts. EDBGP did not inhibit glycolysis (glucose consumption and lactate production). Pre-treatment of lung cancer cells with several antioxidants did not reduce the cytotoxicity of EDBGP, thereby indicating that reactive oxygen species do not participate in the anticancer activity of this compound. Finally, EDBGP was screened against a panel of cancer cells and normal cells from several tissues, including three genetically modified skin fibroblasts with increasing degree of malignancy. Our results suggest that epoxyalkyl galactopyranosides are promising lead compounds for the development of new anticancer agents.

Ice-binding proteins (IBPs) have several functions that permit their hosts to thrive in the presence of ice. The ability of IBPs to control ice growth makes them potential additives in various industries ranging from food storage and cryopreservation to anti-icing systems. For IBPs to be used in commercial applications, however, methods are needed to produce sufficient quantities of high-quality proteins. Here, we describe a new method for IBP purification, termed falling water ice purification (FWIP). The method is based on the affinity of IBPs for ice. A crude IBP solution is allowed to flow continuously over the large chilled vertical surface of a commercial ice machine. The temperature of the surface is lowered gradually until ice crystals are produced, to which the IBPs bind but other solutes do not. As in other ice affinity methods, FWIP does not require molecular tags and is suitable for purifying recombinant IBPs as well as IBPs from natural sources. The advantage of FWIP over other ice affinity methods is that it exploits an ice machine designed to produce large volumes of clear ice daily. This system can be easily scaled up and suits the purification of industrial quantities of IBPs. The FWIP method significantly advances the use of IBPs in research and industry.

Bacterial pathogens have been attributed to poultry housing structure, financial strength, and incessant use of antibiotics, variable seasons and management systems practiced. Variant forms of bacterial pathogens can be detected by recognizing the molecular pattern of the pathogens through an innate immune mechanism such as mannose-binding lectin. Mannose-binding lectin (MBL) possesses an innate pattern recognition molecule that easily sequestered to region of infections and inflammations. This works by attaching itself to antigen surface thus hinders proliferation and disease activity in the host organism. Baker’s method, nephelometric assays technique, Enzyme-Linked Immunosurbent Assay technique, Polymerase Chain Reaction, Deoxyribonucleic Acid typing and other biotechnology related methods are techniques used in detecting and quantifying MBL. Mannose-binding lectin levels in serum can be influenced by age, management systems, feed formulation strategies and seasons. Therefore, knowledge of MBL should be encouraged in all aspect of poultry production, in order to discourage incessant use of drugs at a slight exposure to prevailing bacterial which can help in maximizing cost.

Following the concept of strong interaction, theoretically, fusion of proton seems to increase the binding energy of final atom by 8.8 MeV. Due to Coulombic repulsion, asymmetry effect, pairing effect and other nuclear effects, final atom is forced to choose a little bit of binding energy less than 8.8 MeV and thus it is able to release left over binding energy in the form of internal kinetic energy or external thermal energy. Thus, in cold fusion, heat release to occur, binding energy difference of final atom and base atom seems to be less than 8.8 MeV. Qualitatively, energy released during cold fusion seems to be approximately equal to 8.8 MeV minus the difference of binding energy of final and base atoms. Based on this idea, under normal conditions, for the case of ₂He⁴, fusion of four protons can liberate (35.2-28.3)=6.9 MeV and it is 3.5 times less than the current estimates. Point to be understood is that, lesser the binding energy of final atom, higher the liberated thermal energy and vice versa. With a suitable catalyst and sufficient hydrogen under suitable pressure, if reactor’s temperature is maintained at (1000 to 1500) ⁰C, there seems a lot of scope for a chain reaction of cold fusion in which light isotopes transform to their next stage with increased proton number or mass number and liberate safe and clean heat energy continuously. By arranging 4 to 6 reactors and charging them periodically in tandem, required thermal energy can be produced continuously. In this new direction, by carefully selecting the base isotope and its corresponding catalyst, experiments can be conducted and ground reality of cold fusion can be understood at various temperature and pressure conditions.

Nuclear export of viral mRNAs, is an essential step in the HIV replication cycle. This role is played by a small regulatory protein of HIV-1 called Rev.The N-terminal region of Rev contains an arginine-rich sequence. The arginine-rich motif (ARM) is located between amino acids 38-50 and forms an alpha-helical secondary structure. Expression of the structural proteins of human immunodeficiency virus type 1 requires the direct interaction of multiple copies of the viral Rev protein with its highly structured RNA target sequence, the Rev Response element (RRE). The major viral proteins are not produced if this transport of RNA is stopped. Therefore, knowledge of Rev structure is essential for understanding of its cooperative binding to the RRE, for understanding the mechanism of HIV infection and for the development of antiviral drugs that interfere with Rev’s essential functions and for acknowledgment of good candidate drugs for treatment of AIDS. To understand how REV interact with RRE element of HIV-RNA and its formation of oligomeric complex it is better to characterize the domain wise structure of REV with regard in function of each domain. Due to lack of structural data on Rev no single compound is reported as inhibitor of REV expect antiviral drugs. Identification of a high-affinity RNA-binding site for the human immunodeficiency virus type 1 Rev Protein is much more important. The ARM is a highly specific sequence which allows for the multimerization of Rev and also binding of REV with RNA. Here we are first time exploring the structural characteristics of REV protein both in free form and in complex with RNA at domain function level especially explore the role of ARM motif in REV HIV-1 protein as RNA binding sites by molecular dynamics (MD) simulation and homology modeling studies. Results indicate that the arginine-rich motif (ARM) is crucial in stability of this complex. The residues ARG38, 39, 41, 43, 44, 48, 50, and ASN40 are most interacting with nucleobases of RRE in Crystal structure of Rev and Rev-response-element RNA complex. Our study plays a major role in elaboration of binding of RNA with REV and pave the way for further investigation for therapeutically agent for HIV.

Aptamers are versatile oligonucleotide ligands used for molecular recognition of diverse targets. However, application of aptamers to the field of amyloid β-protein (Aβ) has been limited so far. Aβ is an intrinsically disordered protein that exists in a dynamic conformational equilibrium, presenting time-dependent ensembles of short-lived, metastable structures and assemblies that have been generally difficult to isolate and characterize. Moreover, despite understanding of potential physiological roles of Aβ, this peptide has been linked to the pathogenesis of Alzheimer disease, and its pathogenic roles remain controversial. Accumulated scientific evidence thus far highlights undesirable or nonspecific interactions between selected aptamers and different Aβ assemblies likely due to metastable nature of Aβ or inherent affinity of RNA oligonucleotides to β-sheet-rich fibrillar structures of amyloidogenic proteins. Accordingly, lessons drawn from Aβ–aptamer studies emphasize that purity and uniformity of the protein target and rigorous characterization of aptamers’ specificity are important for realizing and garnering the full potential of aptamers selected for recongizing Aβ or other intrinsically disordered proteins. This review summarizes studies of aptamers selected for recognizing different Aβ assemblies and highlights controversies, difficulties, and limitations of such studies.

Nonalcoholic fatty liver disease (NAFLD) is becoming the leading cause of hepatocellular carcinoma (HCC), liver-related mortality, and liver transplantation. There is reasonable epidemiological cohort data to recommend surveillance of patients with NAFLD based upon the incidence of HCC. The American Gastroenterology Association (AGA) expert review published in 2020 recommend that NAFLD patients with cirrhosis or advanced fibrosis estimated by non-invasive tests (NITs) should consider HCC surveillance. NITs include fibrosis-4 (FIB-4) index, the enhanced liver fibrosis (ELF) test, FibroScan, and MR elastography. The recommended surveillance modality is abdominal ultrasound (US) given that it is cost effective and noninvasive with good sensitivity. However, US is limited in obese patients and those with NAFLD. In NAFLD patients with a high likelihood of having an inadequate US or if US is attempted but inadequate, CT or MRI may be utilized. The GALAD score, consisting of age, gender, AFP, lens culinaris-agglutinin-reactive fraction of AFP (AFP-L3), and protein induced by vitamin K absence or antagonist-II (PIVKA-II), can help to identify high risk of incident HCC in NAFLD patients. Innovative parameters including Mac-2 binding protein glycated isomer , type IV collagen 7S, free apoptosis inhibitor of macrophage, combination of single nucleoside polymorphisms are expected to be established. Considering a large number of NAFLD population, optimal screening tests must meet several criteria including high sensitivity, cost effectiveness and availability.

The antioxidant activity of molecules constitutes an important factor for the regulation of redox homeostasis and reduction of oxidative stress. Cells affected by oxidative stress can undergo genetic alteration, causing structural changes and promoting the onset of chronic diseases, such as cancer. The in silico study performed here was developed to evaluate the antioxidant potential of two molecules, ZINC08706191 (Z91) and ZINC08992920 (Z20), with recognized epithelial anticancer potential. Molecular docking, chemical-quantum calculations and Pearson's correlation were performed. The Z91 and Z20 molecules showed lower binding free energy (ΔG) values for the receptor-ligand interaction than the reference molecules (caffeine – CAF and ascorbic acid – AA), and better results for values of molecular descriptors correlated with ΔG, resulting in a decrease of ΔG. Strong correlations were observed between ΔG values for the five receptors evaluated and ΔG values of the potential epithelial anticancer activity available in literature. These results attest to the significant antioxidant potential of the Z91 and Z20 molecules and their strong relation with the potential epithelial anticancer activity and may be indicated for further analysis in relation to the control of oxidative stress and epithelial anticancer activity.

PHA synthases (PhaC) are grouped into four classes based on the kinetics and mechanisms of reaction. The grouping of PhaC enzymes into four classes is dependent on substrate specificity, according to the preference in forming short chain length (scl) or medium chain length (mcl) polymers: class I, class III, and class IV produce scl-PHAs depending on propionate, butyrate, valerate and hexanoate precursors, while class II phaC synthesize mcl-PHAs based on the alkane (C6 to C14) precursors. PHA synthases of class I, in particular PhaC_Cs from Chromobacterium USM2 and PhaC_Cn/RePhaC1 from Cupriavidus necator/R. eutropha, have been analysed and the crystal structures of the C-domains have been determined. PhaC_Cn/RePhaC1 was also studied by small angle X-ray scattering (SAXS) analysis. Models have been proposed for dimerization, catalysis mechanism, substrate recognition and affinity, product formation and product egress route. The assays based on amino acid substitution by mutagenesis have been useful to validate the hypothesis on the role of amino acids in catalysis and in accommodation of bulky substrates, for the synthesis of PHB co-polymers and medium chain length-PHA polymers with optimized chemical properties.

The SARS (severe acute respiratory syndrome)-CoV (Coronavirus)-2 S(spike)-protein mRNA/cDNA currently being used as vaccines are antigenic but not antigens against SARS-CoV-2, that causes COVID (Coronavirus Disease) -19. Furthermore, the mRNA and cDNA antigenic vaccines also have potentials for homologous as well as heterologous recombination, primarily into the somatic cell DNA of the vaccine recipients. On the contrary, a SARS-CoV-2 RBD-protein antigen, a part of the S-protein, will directly stimulate antibody production against SARS-CoV-2. Hence, a vaccine composed of SARS-CoV-2 RBD-protein as a safer, fast acting, and effective vaccine against SARS-CoV-2 and thus against COVID-19. This is also useful for some immune compromised individuals.

We recently screened a series of new aziridines β-D-galactopyranoside derivatives for selective anticancer activity and identified 2-methyl-2,3-[N-(4-methylbenzenesulfonyl)imino]propyl 2,3-di-O-benzyl-4,6-O-(S)-benzylidene-β-D-galactopyranoside (AzGalp) as the most promising compound. In this article, we explore possible mechanisms involved in the cytotoxicity of this aziridine and evaluate its selective anticancer activity using cancer cells and normal cells from a variety of tissues. Our data show that AzGalp induces DNA damage (detected with the comet assay). Cells deficient in the DNA repair pathway nucleotide excision repair (NER) were hypersensitive to the cytotoxicity of this compound. These results suggest that AzGalp induces bulky DNA adducts, and that cancer cells lacking a functional NER pathway may be particularly vulnerable to the anticancer effects of this aziridine. Several experiments revealed that neither the generation of oxidative stress nor the inhibition of glycolysis played a significant role in the cytotoxicity of AzGalp. The combinations of AzGalp with either oxaliplatin or 5-fluorouracil slightly improved the ability of both anticancer drugs to selectively kill cancer cells. AzGalp also displayed selective cytotoxicity against a panel of malignant cells versus normal cells; the highest selectivity was observed for two acute promyelocytic leukemia cell lines. Additional preclinical studies are necessary to evaluate the anticancer potential of AzGalp.

Emerging paradigms in interferon (IFN) biology suggest a dynamic INF induced interactome that extends through broader Interferon Stimulated Gene (ISG)- induction, which implicates interferon- ISG coordinated cross-talk with mRNA processing, post-translational modification and metabolic processes that underlie pathological (viral, autoimmune and tumor biology) and physiological (stem cell regenerative pathways) processes. INF immune responses can also be triggered by endogenous host-derived molecules that are generated in response to cellular stress or hemostasis imbalance to establish tissue repair and regeneration in first place, however, overactivation or lack of countermeasures can result in host tissue damage. The proteases are integral to viral and tumor pathology, and importantly serine proteases TMPRSS2 and trypsin have been identified as important molecular determinants underlying COVID-19 pathology, and emergence of coronaviruses cultured in vitro, respectively. We propose that pathogen associated proteases can act as novel stress-inducers to facilitate viral- competent immunomodulation. We term it as Protease Induced Transcriptomic/ epi-Transcriptomic Reshaping (PITTR) of host cells to counter cellular stress. We present a novel experimental model and our preliminary findings of trypsin- primed Caco-2 cells (CPT) that result in translational halt comparable to cells grown under serum-starvation conditions (CSS). CPT at escalating trypsin concentration (CPT- EC) induce upregulation of selective proteins that majorly map to ribosomal, RNA transport, and spliceosome ribonucleoproteins (RNPs). The inclusion of proinflammatory IL1-b to CPT (CPT- IL) resulted in global overexpression of proteins comparable to Caco-2 cells cultured in growth-factor rich serum conditions (CFBS), indicating a likely de-repression of trypsin- induced translational halt. Caco-2 cells display abortive interferon proteome under differential trypsin conditions (CPT, CPT-EC and CPT-IL), which is marked by complete lack of INF generation despite induction of intermediate ISGs, suggestive of protease (trypsin)- dependent regulation of INF response. Viruses regulate the proteome of stress granules (SGs) that are induced to cope transient translational halt as a central adaptive response to pathogen induced cellular stress. The integral components of SGs include non-translating mRNA, ribonucleoproteins (RNPs) and RNA binding proteins (RBPs), which together form biological condensates through a biophysical process involving weak electrostatic interactions through intrinsically disordered regions in RBPs resulting in liquid- liquid phase separation. We compared the CPT- EC proteome to the Mammalian Stress Granules Proteome (MSGP) database to explore potential RBPs that could possibly regulate INF response (and could act as potential anti-viral targets). Notably, differentially upregulated RNPs and potential RBPs from ISG family including ADAR and PRKRA, and RNA helicases implicated in viral pathogenesis were found to be upregulated in the CPT- EC proteome further strengthening the role of proteases (trypsin) in regulating INF pathways independent of the pathogen. We propose that the supplementation of viable SARS-CoV-2 viral loads to trypsin- primed host cells could recapitulate an infectious disease model, which may closely phenocopy pathogen- driven inflammation and signaling events. Based on the global downregulation of seven SERPINS (serine protease inhibitors) linked to thromboinflammation in our LCMS profiling data, we support the candidature of serine protease inhibitors for protease mediated viral pathologies. COVID-19 is increasingly linked to coagulopathy and resemblance to Neutrophil Extracellular Trap (NET) related thromboinflammatory features; SERPIN A1AT (alpha 1 antitrypsin) being a potent neutrophil- elastase inhibitor and a negative regulator of coagulation complement pathway may be a promising candidate for establishing hemostasis rebalancing in COVID-19 pathology.

We have previously reported that 60% sucrose diet-fed ChREBP knockout mice (KO) showed body weight loss resulting in lethality. We aimed to elucidate whether sucrose and fructose metabolism are impaired in KO. Wild type mice (WT) and KO were fed a diet containing 30% sucrose with/without 0.08% miglitol, an α-glucosidase inhibitor, and these effects on phenotypes were tested. Furthermore, we compared metabolic changes of oral and peritoneal fructose injection. Thirty percent sucrose diet feeding did not affect phenotypes in KO. However, miglitol induced lethality in 30% sucrose-fed KO. Thirty percent sucrose plus miglitol diet-fed KO showed increased cecal contents, increased fecal lactate contents, increased growth of lactobacillales and Bifidobacterium and decreased growth of clostridium cluster XIVa. ChREBP gene deletion suppressed the mRNA levels of sucrose and fructose related genes. Next, oral fructose injection did not affect plasma glucose levels and liver fructose contents; however, intestinal sucrose and fructose related mRNA levels were increased only in WT. In contrast, peritoneal fructose injection increased plasma glucose levels in both mice; however, the hepatic fructose content in KO was much higher owing to decreased hepatic Khk mRNA expression. Taken together, KO showed sucrose intolerance and fructose malabsorption owing to decreased gene expression.

Both SARS-CoV-2 and SARS-CoV initially appeared in China and spread to other parts of the world. SARS-CoV-2 has generated a COVID-19 pandemic causing more than 6 million human deaths worldwide while the SARS outbreak quickly ended in six months with a global total of 774 reported deaths. One of the factors contributing to this stunning difference in the outcome between these two outbreaks is the inaccuracy of the RT-qPCR tests for SARS-CoV-2, which generated a large number of false-negative and false-positive test results that have misled patient management and public health policy-makers. This article presented Sanger sequencing evidence to show that the RT-PCR diagnostic protocol established in 2003 for SARS-CoV can in fact detect SARS-CoV-2 accurately due to the well-known nonspecific PCR amplification of DNAs with similar sequences. Using nested RT-PCR followed by Sanger sequencing to retest 50 patient samples collected in January, 2022 and sold as RT-qPCR positive reference confirmed 21 (42%) were false-positive. Although the other 29 positive isolates were categorized as Omicron variant by partial sequencing of the N gene, and the RBD and the NTD of the S gene, 9 (31%) showed focal to complete sequencing failure in the S gene segments due to multi-allelic SNPs. During the course of the study, an Omicron variant isolate containing a BA.1 NTD and a BA.2 RBD in its S gene was also detected. Routine partial S gene sequencing of all PCR-positive samples can timely discover multi-allelic SNPs and viral recombination in the circulating variants for investigation of their impacts on vaccine efficacies, therapeutics and diagnostics.

Single nucleotide polymorphisms (SNP) are associated with diseases and drug response variabilities in humans. Elucidating the damaging and disease-associated SNPs using wet-laboratory approaches can be challenging and resource-demanding due to the large number of SNPs in the human genome. Due to the growth in the field of computational biology and bioinformatics, algorithms have been developed to help screen and filter out the most deleterious SNPs that are worth considering for wet-laboratory studies. Here we review the existing in-silico based methods used to predict and characterize the effects of SNPs on protein structure and function. This cutting-edge approach will facilitate the search for novel therapeutics, help understand the etiology of diseases and fast-track the personalized medicine agenda.

Euglena gracilis is an alga of great biotechnological interest and extensive metabolic capacity, able to make high levels of bioactive compounds, such as polyunsaturated fatty acids, vitamins and β-glucan. Previous work has shown that Euglena expresses a wide range of carbohydrate-active enzymes, suggesting an unexpectedly high capacity for the synthesis of complex carbohydrates for a single-celled organism. Here, we present an analysis of some of the carbohydrates synthesised by Euglena gracilis. Analysis of the sugar nucleotide pool showed that there are the substrates necessary for synthesis of complex polysaccharides, including the unusual sugar galactofuranose. Lectin- and antibody-based profiling of whole cells and extracted carbohydrates revealed a complex galactan, xylan and aminosugar based surface. Protein N-glycan profiling, however, indicated that just simple high mannose-type glycans are present and that they are partially modified with putative aminoethylphosphonate moieties. Together, these data indicate that Euglena possesses a complex glycan surface, unrelated to plant cell walls, while its protein glycosylation is simple. Taken together, these findings suggest that Euglena gracilis may lend itself to the production of pharmaceutical glycoproteins.

New flaxseed cultivars differing in seed mucilage content (MC) with low hull content (HC) represent an attractive option to simultaneously target the food and feed markets. Here, a genome-wide association study (GWAS) was conducted for MC and HC in 200 diverse flaxseed accessions genotyped with 1.7 million SNP markers. The data obtained for MC and HC indicated a broad phenotypic variation and high (~70%) and a moderate (~49%) narrow sense heritability, respectively. MC and HC did not differ statistically between fiber and oil morphotypes, but yellow-seeded accessions had 2.7% less HC than brown-seeded ones. The genome wide linkage disequilibrium (LD) decayed to r2 = 0.1 at a physical distance of ~100 Kb. Seven and four QTL were identified for MC and HC, respectively. Promising candidate genes included Linum usitatissimum orthologs of the Arabidopsis thaliana genes TRANSPARENT TESTA 8, SUBTILISIN-LIKE SERINE PROTEASE, GALACTUROSYL TRANSFERASE-LIKE 5, MUCILAGE-MODIFIED 4, AGAMOUS-LIKE MADS-BOX PROTEIN AGL62, GLYCOSYL HYDROLASE FAMILY 17 and UDP-GLUCOSE FLAVONOL 3-O-GLUCOSYLTRANSFERASE that have been shown to play a role in mucilage synthesis and release, seed coat development and anthocyanin biosynthesis in A. thaliana were identified. The favorable alleles will be useful in flaxseed breeding towards the goal of achieving the ideal flaxseed cultivars for food and feed by genomic-based breeding.

In this work, kinetics of the reaction between 5,10,15,20-tetrakis(N-methylpyridium-4-yl)porphyrin and Ni2+ species were investigated in aqueous solution at 25 ±1 ºC in I = 0.10 M (NaNO3). Speciation of Ni2+ was carried out in I = 0.10 M (NaNO3) in order to provide the distribution of the Ni2+ species with different solution pH. The experimental data have been compared with the speciation diagram constructed from the values of hydrolysis constants of Ni2+ ion. Speciation data showed that the hexaaquanickel(II), [Ni(H2O)6]2+, ions take place in hydrolysis reactions through formation of [Ni(OH2)6-n(OH)n]2-n species with solution pH. Based on the speciation of Ni2+ and pH dependent rate constants, rate expression can be written as: d[Ni(TMPyP)4+]/dt = (k1[Ni2+(aq)] + k2[Ni(OH)+(aq)] + k3[Ni(OH)2o(aq)] + k4[Ni(OH)3-(aq)])[H2TMPyP4+], where k1, k2, k3 and k4 were found to be k1 = (0.62 ± 0.22) × 10-2; k2 = (3.60 ± 0.40) × 10-2; k3 = (2.09 ± 0.52) × 10-2, k4 = (0.53 ± 0.04) × 10-2 M-1s-1 at 25 ±1 °C, respectively. Kinetic results showed that monohydroxo, [Ni(H2O)5(OH)]+, is the most reactive among the [Ni(OH2)6-n(OH)n]2-n species. The enhanced reactivity has been ascribed to the formation of hydrogen bonding between oxygen atom of hydroxyl group of the [Ni(H2O)5(OH)]+ species and the pyrrolic hydrogen atom of the [H2TMPyP]4+. The rate of formation of [Ni(II)TMPyP]4+ complex was to be 3.99 × 10-2 M-1s-1 in I = 0.10 M, NaNO3 (25 ± 1 ºC). Ionic strength effect on the reaction rate is suggested that the net charge of the tetracationic porphyrin is to be +3.6 on the basis of Brønsted-Bjerrum equation. The UV-Vis and fluorescence data revealed that [Ni(II)TMPyP]4+ and H2(TMPyP)4+ interact with DNA, and UV-Vis results suggest that Ni(II)-porphyrin and free base porphyrin interact with DNA via outside binding with self-stacking and intercalation, respectively. Mechanism of kinetics of formation of the [Ni(II)TMPyP]4+ complex in aqueous medium is discussed. An investigation of application of the [Ni(II)TMPyP]4+ complex along with other metalloporphyrins such as Zn2+-, Ru2+-, Pt2+-, [Au(III)TMPyP]5+ as anti-COVID-19 agents is now in progress under international collaboration.

Physics behind collimated highly directional nature of lasers, and factors that keep the seven coloured waves that form white light together during their journey from Sun to Earth, in the face of the natural disruptive forces, is not fully understood. Energy levels were measured, in terms of alterations in induced current and voltage, in beams from a red laser, white LED light and the Sunlight before and during their disruption by diffusers (frosted glass for the lasers) and diffractors (diffraction grating for the white light) using a photovoltaic solar cell panel attached to a digital multimeter. Results show that disruption of the beams results in release of extra energy named as ‘Latent Light Binding’ Energy’. It is hypothesized that the ‘binding’ energy keeps laser waves firmly bound together both end-on and side-on enabling laser beams to travel long distances in collimated manner. Likewise, the 7 coloured waves that constitute white light are kept together, probably side-on, in their journey from the Sun to the Earth. The observation that diffraction of sunbeam is associated with increased power generation provides a new lead to improve harnessing of solar energy, where, currently, the focus is mainly on improving efficiency of photovoltaic cell.

Identifying anti-spike antibodies that exhibit strong neutralizing activity against current dominant circulating variants and antibodies that are escaped by these variants have important implications in the development of therapeutic and diagnostic solutions as well as in improving understanding of the humoral response to SARS-CoV-2 infection. We characterized seven anti-RBD monoclonal antibodies for their binding activity, pairing capability and neutralization activity to SARS-CoV-2 and three variant RBDs (UK, SA and BR P.1) via lateral flow immunoassays. The results allowed us to group these antibodies into three distinct epitope bins. Our studies showed that two antibodies had broadly potent neutralizing activity against SARS-CoV-2 and these variant RBDs and that one antibody did not neutralize the SA and BR P.1 RBDs. The antibody escaped by the SA and BR P.1 RBDs retained binding activity to SA and BR P.1 RBDs but was unable to induce neutralization. Further, we demonstrated that the lateral flow immunoassay can be a rapid and effective tool for antibody characterization, including epitope classification and antibody neutralization kinetics. From these studies, the potential contributions of the mutations (N501Y, E484K and K417N/T) contained in these variants’ RBDs on antibody pairing capability, neutralization activity and therapeutic antibody targeting strategy are discussed.

The regulation of hepatic very-low-density lipoprotein (VLDL) secretion plays an important role in the pathogenesis of dyslipidemia and fatty liver diseases. VLDL is controlled by hepatic microsomal triglyceride transfer protein (MTTP). Mttp is regulated by carbohydrate response element binding protein (ChREBP) and small heterodimer partner (SHP). However, it is unclear whether both coordinately regulate Mttp expression and VLDL secretion. Here, adenoviral overexpression of ChREBP and SHP in rat primary hepatocytes induced and suppressed Mttp mRNA, respectively. However, Mttp induction by ChREBP was much more potent than suppression by SHP. Promoter assays of Mttp and the liver type pyruvate kinase gene revealed that SHP and ChREBP did not affect the transactivity of each other. Mttp mRNA and protein levels of Shp^/– mice were similar to those of wild-type; however, those of Chrebp^–/–Shp^–/– and Chrebp^–/– mice were much lower. Consistent with this, the VLDL particle number and VLDL secretion rates in Shp^–/–- mice were similar to wild-type, but were much lower in Chrebp^–/– and Chrebp^–/–Shp^–/–- mice. These findings suggested that ChREBP rather than SHP regulates VLDL secretion and that ChREBP and SHP do not affect the transactivities of each other.

Muscle is one of the most critical organs for mammals, which governs multiple movement and physiological functions. Circular RNA (circRNA) is a kind of novel endogenous RNA without 5'-Caps and 3'-poly(A) structures formed by pre-mRNA's back-splicing. RNA binding proteins (RBPs) control the production and degradation of circRNA, help nucleus-cytoplasm transport and locate circRNA, and regulate circRNA translation. Therefore, circRNAs and the chaperoned RBPs play critical roles in muscle growth, development, and disease progression. In this review, we systematically characterize the possible molecular mechanism of circRNA-protein interactions. Also, we summarize the latest researches on circRNA-protein interactions in muscle development and diseases. Besides, we provide several valid prediction methods and experimental verification approaches. Our review reveals the importance of circRNAs and their protein chaperones and provides a reference for further study in this field.

HIF-1α is a master regulator of oxygen homeostasis involved in different stages of cancer development. Thus, HIF-1α inhibition represents an interesting target for anti-cancer therapy. It was recently shown that HIF-1α interaction with NQO1 inhibits its proteasomal degradation, thus suggesting that targeting the stability of NQO1 could led to destabilization of HIF-1α as a therapeutic approach. Since the molecular interactions of NQO1 with HIF-1α are beginning to be unraveled, we review here our current knowledge on the intracellular functions and stability of NQO1, its pharmacological modulation by small ligands, and the molecular determinants of its roles as a chaperone of many different proteins including cancer-associated factors such as p53 and p73α. This knowledge is then discussed in the context of potentially targeting the intracellular stability of HIF-1α by acting on its chaperone, NQO1. This could result in novel anti-cancer therapies.

Ubiquitin binding domains (UBDs) are modular elements that bind non-covalently to ubiquitin and act as downstream effectors and amplifiers of the ubiquitination signal. With few exceptions, UBDs recognize the hydrophobic path centered on Ile44 (Leu-8, Ile-44, Val-70). Nevertheless, a variety of different orientations, which can be attributed to specific contacts between each UBD and surface residues surrounding the hydrophobic patch, specify how each class of UBD recognizes ubiquitin. Here, we describe the structure of a novel ubiquitin-binding domain that we identified in NEDD4 binding protein 1 (N4BP1). By performing protein sequence analysis, mutagenesis and NMR spectroscopy of the ¹⁵N isotopically labelled protein, we demonstrate that a Phe-Pro motif in N4BP1 recognizes the canonical hydrophobic patch of ubiquitin. This recognition mode resembles the molecular mechanism evolved in the CUE (Coupling of ubiquitin conjugation to ER degradation) domain family, where an invariant proline, usually following a phenylalanine, is required for binding to ubiquitin. Interestingly, the UBD of N4BP1 is evolutionary related to CUBAN (Cullin binding domain associating with NEDD8) (40% identity and 47% similarity), a protein module that also recognizes the ubiquitin-like NEDD8, which is the closest relative of ubiquitin (58% identity and 80% similarity). By performing circular dichroism and ¹⁵N NMR chemical shift perturbation of N4BP1 in complex with ubiquitin, we demonstrate that the UBD of N4BP1 lacks the NEDD8 binding properties observed in CUBAN and it recognizes the Ile44-centered patch of ubiquitin through a dedicated binding site, which share some of the features observed in the CUE domain family. Moreover, we show that, in addition to mediating the interaction with ubiquitin and ubiquitinated substrates, both the CUBAN and the UBD of N4BP1 are poly-ubiquitinated in cells. This modification is dependent on the presence of a functional domain. We believe that the structural and functional characterization of this novel UBD will allow a deeper understanding of the molecular mechanisms governing N4BP1 function, while at the same time providing a valuable tool for clarifying how the discrimination between ubiquitin and the highly related NEDD8 is achieved.

A precise understanding of the genes and associated genomes of SARS-CoV2 is important for various reasons such as discovering origin of the virus and virulence and so on. A thorough descriptive understanding of the SARS-CoV2 genomes and other coronavirus of the beta-coronavirus genus is primarily important. In this article, a set of ten genomes of four CoVs and their associated genes are considered for this present study. A spatial representations of nucleotide bases including purine-pyrimidine representations of the different genes of the corresponding genomes are quantified using Hurst exponent, Shannon entropy and density estimation of different nucleotides including GC content, in order to draw a comparison and contrast among the ten genomes of different types of CoVs which include MERS, SARS-CoV, HKU1 (Human Coronavirus) and associated their genes.

C-C Chemokine receptor type 5 (CCR5) is expressed on the CD4 T cell surface where CXCR4 and CCR5 expressions are controlled differently during the activation of T cells and with the binding of interleukin type-2 (IL-2). IL-2 can upregulate CCR1 and CCR2 in CD45R01 T cells and increase the T lymphocyte chemotaxis toward CC-chemokines. CD4+ T cells are either apart of the T helper 1 or Th1 lymphocytes that release interferon gamma (IFNγ) and lymphotoxin that provide cellular immunity to internal pathogens and T helper cells type 2 (Th2), which secrete interleukins 4 and 5 (IL-4 and IL-5). IL-4 and IL-5 cause an allergic and humoral immune response to parasites. Th2 lymphocytes use CCR3 chemokine receptors. CCR5 and CXCR3 chemokine receptors are specific for CD4+ Th1 and Th2 lymphocytes. CCR5Δ32 is a 32-base-pair deletion of the CCR5 gene. CCR5 is a co-receptor for the entrance of the human immunodeficiency virus-1 (HIV-1). CCR5Δ32 creates a malfunction of the CCR5 protein that can prevent HIV-1 infection. However, the CCR5Δ32 32-base-pair deletion is not prevalent and predominant in many populations worldwide and there also exist more genetic variations of CCR5 known as CCR5-SNPs. An alternative polymorphism was identified based on the CCR5 gene that was identified as a A to G (A/G) point mutation. This point mutation is located at the 59029 locus on the promoter that lowers the expression rate of CCR5. Gurdol et al. found activity at the promoter of CCR5/590029G was 45% lesser than the CCR5/59029A. The genetic variations of CCR5 SNPs also include: 2459G>SNP of CCR5, C101X, CCR5 gene position at -2273, and the A to G SNP mutation found in two South African blacks. Therefore, more studies are needed to find more and varied chemokine polymorphisms that are present in many diverse populations in the world. The aim of this literature review is to describe the immense impact of CCR5 SNP mutations on viral infection susceptibility, on the pathogenesis of chronic conditions, to endorse the increased discovery of more novel CCR5 SNPs, and to show the significant potential of anti-CCR5 therapies to treat multiple diseased conditions.

Remdesivir (RDV) has garnered much hope for its moderate anti-COVID-19 effects, but its limited amelioration of survival in hospitalized patient causes a huge controversy over the applicability of RDV to COVID-19 treatment. Developing strategies to improve its antivirus efficacy is urgently required. As anticipated, RDV exhibits similar behavior with other nucleotide analogs to disrupt the metabolism of natural endogenous ribonucleotides (RNs) and deoxyribonucleotides (dRNs). Alterations in endogenous RNs and dRNs play a critical role in virus replication as well as other key cellular functions. Thus elucidation of the disturbances of RDV on RNs and dRNs could help to understand its exact mechanism of action. Here, the metabolic profiling determined by liquid chromatography–mass spectrometry method showed a general increase in the abundance of nucleotides and a more than 2-fold increase for specific nucleotides. However, the variation of pyrimidine ribonucleotides was relative slight or even contrary, resulting in obvious imbalance between purine and pyrimidine ribonucleotides, which implied the obstacle of RDV to pyrimidine synthesis and could further block the transcription and replication of viral RNA. Additionally, the extreme disequilibrium between cytidine triphosphate (CTP) and cytidine monophosphate might result from the inhibition of CTP synthase and provide a metabolic target for the treatment of COVID-19 infection. Since nucleotides metabolism pathways are vulnerable to nucleotide analogues and are liable to be the regulation targets, it is promising to enhance the efficacy of RDV through co-administration with CTP synthase inhibitors or de novo pyrimidine synthesis inhibitors to exacerbate the imbalance of nucleotide pools.

Immunosensors have been widely developed to use antibodies to detect a pathogen of interest; it is interesting to look at the effect of nonspecific antibody binding to E. coli using electrochemical methods. IgG antibody not specific to E. coli O157:H7 was crosslinked onto a screen-printed carbon electrode. The presence of E. coli at 4, 4 × 10², 4 × 10⁵, and 4 × 10⁸ CFU/mL on the electrode surface was detected via linear sweep voltammetry (LSV) and electrochemical impedance spectroscopy (EIS). Current transfer at both electrodes was reduced as the concentration of bacteria increased; however, the calibration of number of cells to decreased current was nonlinear for IgG-modified electrodes. The nonlinearity is confirmed by EIS measurements which showed highest impedance at 4 CFU/mL E. coli when impedance should be the lowest. FESEM images showed higher binding of cells when IgG is present compared to electrodes with reduced graphene oxide (rGO) alone. Electrodes with rGO alone show less attachment of E. coli, with EIS showing a linear calibration profile, while LSV shows not much difference in current values for all concentrations aside from the highest concentration. These results suggest that nonspecific binding can provide false signals in electrochemical measurements, and it is crucial to provide proper controls.

FRAP technique have been used for decades to measure movements of molecules in 2D. Data obtained by FRAP experiments in cell plasma membranes are assumed to be described through means of two parameters, a diffusion coefficient D (as defined in a pure Brownian model) and a mobile fraction M. Nevertheless, it has also been shown that recoveries can be nicely fit using anomalous sub-diffusion. FRAP at variable radii has been developed using the Brownian diffusion model to access geometrical characteristics of the surrounding landscape of the molecule. Here we performed numerical simulations of continuous time random walk (CTRW) anomalous subdiffusion and interpreted them in the context of variable radii FRAP. These simulations were compared to experimental data obtained at variable radii on living cells using the PH domain of the membrane binding protein EFA6 (exchange factor for ARF6, a small G protein). This protein domain is an excellent candidate to explore the structure of the interface between cytosol and plasma membrane in cells. By direct comparison of our numerical simulations to the experiments, we show that this protein does not exhibit anomalous diffusion in BHK cells. The non Brownian PH-EFA6 dynamics observed here is more related to spatial heterogeneities such as cytoskeleton fences effects.

We review the basics of nuclear binding energy scheme assumed to be associated with the existence of a new strong elementary charge associated with square root of reciprocal of the strong coupling constant.

A genome-wide association study (GWAS) was performed on a set of 260 lines which belong to three different bi-parental flax mapping populations. These lines were sequenced to an averaged genome coverage of 19× using the Illumina Hi-Seq platform. Phenotypic data for 11 seed yield and oil quality traits were collected in eight year/location environments. A total of 17,288 single nucleotide polymorphisms were identified, which explained more than 80% of the phenotypic variation for days to maturity (DTM), iodine value (IOD), palmitic (PAL), stearic, linoleic (LIO) and linolenic (LIN) acid contents. Twenty-three unique genomic regions associated with 33 QTL for the studied traits were detected, thereby validating four genomic regions previously identified. The 33 QTL explained 48-73% of the phenotypic variation for oil content, IOD, PAL, LIO and LIN but only 8-14% for plant height, DTM and seed yield. A genome-wide selective sweep scan for selection signatures detected 114 genomic regions that accounted for 7.82% of the flax pseudomolecule and overlapped with the 11 GWAS-detected genomic regions associated with 18 QTL for 11 traits. The results demonstrate the utility of GWAS combined with selection signatures for dissection of the genetic structure of traits and for pinpointing genomic regions for breeding improvement.

Hypercholesterolemia accelerates atherosclerosis, and extensive research has been undertaken to ameliorate this abnormality. Plant sterols have been shown to inhibit cholesterol absorption and lower plasma cholesterol level since the 1950s. This ingredient has recently been reappraised as a food additive that can be taken daily in a preclinical period to prevent hypercholesterolemia, considering that cardiovascular-related diseases are the top cause of death globally even with clinical interventions. Intestinal cholesterol handling is still elusive, making it difficult to clarify the mechanism for plant sterol-mediated inhibition. Notably, although the small intestine absorbs cholesterol, it is also the organ that excretes it abundantly, via trans-intestinal cholesterol efflux (TICE). In this review, we show a model where the brush border membrane (BBM) of intestinal epithelial cells stands as the dividing ridge for cholesterol fluxes, making cholesterol absorption and TICE inversely correlated. With this model, we tried to explain the plant sterol-mediated inhibitory mechanism. As well as cholesterol, plant sterols diffuse into the BBM but are effluxed back to the lumen rapidly. We propose that repeated plant sterol shuttling between the BBM and lumen promotes cholesterol efflux, and plant sterol in the BBM may disturb the trafficking machineries that transport cholesterol to the cell interior.

Matrix Metaloproteinase-2 (MMP-2) is an extracellular Zn²⁺ protease specific to type I and IV collagens. Its expression is associated with several inflammatory, degenerative, and malignant diseases. Conformational properties, domain movements, and interactions between MMP-2 and its associated metal ions were characterized using a 1.0 µs molecular dynamics simulation. Dihedral principle component analysis revealed 10 families of conformations with the greatest degree of variability occurring in the link region connecting the catalytic and hemopexin domains. Dynamics cross correlation analysis indicated domain movements corresponding to opening and closing of the hemopexin domain in relation to the fibronectin and catalytic domains facilitated by the link region. Interaction energies were calculated using the MMPBSA-interaction entropy analysis method and revealed strong binding energies for the catalytic Zn²⁺ ion 1, Ca²⁺ ion 1, and Ca²⁺ ion 3 with significant conformational stability at the binding sites of Zn²⁺ ion 1 and Ca²⁺ ion 1. Ca²⁺ ion 2 diffuses freely away from its crystallographically defined binding site. Zn²⁺ ion 2 plays a minor role in conformational stability of the catalytic domain while Ca²⁺ ion 3 is strongly attracted to the highly electronegative sidechains of the Asp residues around the central β-sheet core of the hemopexin domain.

Aflatoxins in feeds cause great health hazards to animals and in advance, to human. Potential of crude clays designated AC, KC, CC and MC and ashes VA and RA were evaluated for their capacity to adsorb aflatoxins B₁ (AFB₁), B₂ (AFB₂), G₁ (AFG₁) and G₂ (AFG₂) relative to a commercial binder Mycobinder^R (Evonik Industries AG) using in-vitro technique. On average, CC, VA, KC, MC, AC, RA and Mycobind^R adsorbed 39.9%, 51.3%, 61.5%, 62.0%, 72.6%, 84.7% and 98.1% of the total aflatoxins in buffered solution, respectively. The capacity of AC and RA was statistically (p<0.05) better in binding aflatoxins next to Mycobind^R. Capacity of the TBM and Mycobind^R to bind aflatoxins, seemed to follow the trend of their cation exchange capacity (CEC). The CEC (meq/100g) of CC, MC, KC, VA, AC, RA and Mycobind^R were 7.0, 15.4, 18.8, 25.4, 27.2, 27.2 and 38.9, respectively. On average 96.3%, 42.7%, 80.8% and 32.1% of AFB1, AFB2, AFG1 and AFG2 were adsorbed, respectively. Binding capacity of the clays and ashes relative to Mycobind^R was about 100% for AC and RA, 50% for KC, MC and VA and 33.3% for CC. The AC and RA seem to be promising resources in binding aflatoxins in solution.

Study of RNA-protein interactions and identification of RNA targets are among the key aspects of understanding the RNA biology. Currently, various methods are available to investigate these interactions, in particular, RNA pulldown assay. In the present paper, a method based on the HaloTag technology is presented that is called Halo-RPD (HaloTag RNA PullDown). The proposed protocol uses plants with stable fusion protein expression and the MagneBeads magnetic beads to capture RNA-protein complexes directly from the cytoplasmic lysate of transgenic A. thaliana plants. The key stages described in the paper are as follows: 1) preparation of the magnetic beads 2) tissue homogenization and collection of control samples 3) precipitation and wash of RNA-protein complexes; 4) evaluation of protein binding efficacy; 5) RNA isolation; 6) analysis of the obtained RNA. Recommendations for better NGS assay designs are provided.

Structural and biochemical studies have recently revealed a range of rationally engineered nanobodies with efficient neutralizing capacity against SARS-CoV-2 virus and resilience against mutational escape. In this work, we combined atomistic simulations and conformational dynamics analysis with the ensemble-based mutational profiling of binding interactions for a diverse panel of SARS-CoV-2 spike complexes with nanobodies. Using this computational toolkit, we identified dynamic signatures and binding affinity fingerprints for the SARS-CoV-2 spike protein complexes with nanobodies Nb6 and Nb20, VHH E, a pair combination VHH E+U, a biparatopic nanobody VHH VE, and a combination of CC12.3 antibody and VHH V/W nanobodies. Through ensemble-based deep mutational profiling of stability and binding affinities, we identify critical hotspots and characterize molecular mechanisms of SARS-CoV-2 spike protein binding with single ultra-potent nanobodies, nanobody cocktails and biparatopic nanobodies. By quantifying dynamic and energetic determinants of the SARS-CoV-2 S binding with nanobodies, we also examine the effects of circulating variants and escaping mutations. We found that mutational escape mechanisms may be controlled through structurally and energetically adaptable binding hotspots located in the host receptor-accessible binding epitope that are dynamically coupled to the stability centers in the distant epitope targeted by VHH U/V/W nanobodies. The results of this study suggested a mechanism in which through cooperative dynamic changes, nanobody combinations and biparatopic nanobody can modulate the global protein response and induce the increased resilience to common escape mutants.

Exogenous RNA comprises the genetic material associated with several diseases which require immediate treatment, and thus mechanisms to hinder intracellular translation and reproduction of RNA viral agents are of great importance. Applying recent developments from this lab in methods relating to the interaction of DNA with steroid hormones, cyclic compounds are presented for intermolecular binding to nucleic acids. The requirements to achieve binding with RNA nucleotide pairs are described, which involve at a minimum functional elements positioned to interact with the lateral phosphate groups for each of the RNA strands through coupling with a positively charged ion, such as Mg2+, Ca2+, or Zn2+ ions; and an intermolecular hydrogen bond with the oxygen element of uracil at the carbon two location. Additional features of the binding molecules are examined for enhancements and differentiation in binding capability and include aromatic groups that have both a structural role of steric hindrance and a functional role to stabilize the binding mechanisms. Several categories of cyclic compounds are associated to have specific binding capabilities, and the interaction of these structures with potential receptor molecules are evaluated for assessment in delivery and binding of the compound to nucleic acids.

1) Background: We tested whether AHR activation induces DNA damage, whether polymorphisms in genes related to risk of Non-Hodgkin lymphoma are associated with DNA damage, and whether the two conditions do interact with each other. 2) Methods: Our study population included 36 subjects, randomly selected among the population controls participating in a case-control study on lymphoma in Sardinia, Italy, who donated a blood sample. We investigated 47 single nucleotide polymorphisms (SNPs) previously reported to convey risk of lymphoma; the Dual-Glo® Luciferase Assay System to detect activation of the aryl hydrocarbon receptor (AhR) by the serum of study subjects; and the COMET Assay to detect DNA damage. 3) Results: Activation of the aryl hydrocarbon receptor did not increase DNA damage in our study population. On the other hand, the mutant allele (G) of rs1056932/BCL6 increased the occurrence of DNA damage (p = 0.045); such association was confirmed among AhR negative, but not AhR positive subjects. 4) Conclusions: We observed excess DNA damage associated with a gene polymorphism, namely rs1056932/ BCL6, previously reported in association with risk of lymphoma. No increase in DNA damage was associated with AhR activation per se, nor with the other gene polymorphisms we investigated.

Quantitative trait loci (QTL) are genomic regions associated with phenotype variation of quantitative traits in a population. To date, a total of 267 QTL for 29 quantitative traits have been reported in 13 studies on flax. Of these, 200 QTL from 12 studies were identified based on genetic maps, scaffold sequences, or pre-released chromosome-scale pseudomolecules. Molecular markers for QTL identification differed across studies but were mainly based on simple sequence repeat (SSR) or single nucleotide polymorphism (SNP) markers. This article provides methods with software tools and database files to uniquely map SSR and SNP markers from different references onto the recently released chromosome-scale pseudomolecules. Using these methods, 195 QTL were successfully sorted onto the 15 flax chromosomes and grouped into 133 co-located QTL clusters. Mapping of QTL from different studies to the same reference enables comparisons and facilitates genome-wide QTL analysis, candidate gene scanning, and breeding applications.

The rapid expansion of transcriptomics from increased affordability of next-generation sequencing (NGS) technologies generates rocketing amounts of gene expression data across biology and medicine, and notably in cancer research. Concomitantly, many bioinformatics tools were developed to streamline gene expression analysis and quantification. We tested the concordance of NGS RNA sequencing (RNA-seq) analysis outcomes between the two predominant programs for reads alignment, HISAT2 and STAR, and the two most popular programs for quantifying gene expression in NGS experiments, edgeR and DESeq2, using RNA-seq data from a series of breast cancer progression specimens, which include histologically confirmed normal, early neoplasia, ductal carcinoma in situ and infiltrating ductal carcinoma samples microdissected from formalin fixed, paraffin embedded (FFPE) breast tissue blocks. We identified significant differences in aligners’ performance: HISAT2 was prone to misalign reads to retrogene genomic loci, STAR generated more precise alignments, especially for early neoplasia samples. edgeR and DESeq2 produced similar lists of differentially expressed genes in stage comparisons, with edgeR producing more conservative, though shorter, lists of genes. Albeit, Gene Ontology (GO) enrichment analysis revealed no skewness in significant GO categories identified among differentially expressed genes by edgeR vs DESeq2. As transcriptome analysis of archived FFPE samples becomes a vanguard of precision medicine, identification and fine-tuning of bioinformatics tools becomes critical for clinical research. Our results indicate that STAR and edgeR are well-suited tools for differential gene expression analysis from FFPE samples.

Porcine circovirus type 2 (PCV2) is a small non-enveloped DNA virus that causes swine immunosuppression by inducing apoptosis in lymphocytes. The ORF3 protein plays a major role in PCV2-induced apoptosis in porcine kidney cells, but there is little information regarding this protein in PCV2-infected lymphocytes. In this study, hybridoma screening and epitope mapping were determined by using an indirect ELISA. The mAb 7D3 against ORF3 peptide (residues 35–65) of PCV2 were generated in this study. In vivo situation, the mAb 7D3 recognized ORF3 protein existed in PCV2-infected apoptotic porcine PBMCs. It is noteworthy that thimerosal interfered with the binding of mAb 7D3 to epitope and it was diminished by adding cysteine. Additionally, thimerosal interacting with cysteine-containing peptide was demonstrated by the PTI assay. Furthermore, thimerosal specifically interacted with the antigen-binding sites of mAb 7D3. This study suggested that thimerosal blockade the occlusion of the antigen-binding sites of mAb 7D3 to bind ORF3 peptide (residues 35–65) via thimerosal interacting with cysteine residues which should be located within the antigen-binding sites of mAb 7D3. Overall, the mAb 7D3 has been characterized and it will be a valuables tool in future studies of ORF3 function and the wider mechanism of cell apoptosis caused by PCV2 infection. Similarly, these techniques will be useful for applications in detecting thimerosal too.

Antisense technologies consist of the utilization of oligonucleotides or oligonucleotide analogs to interfere with undesirable biological processes, commonly through inhibition of expression of selected genes. This field holds a lot of promise for the treatment of a very diverse group of diseases including viral and bacterial infections, genetic disorders, and cancer. To date, drugs approved for utilization in clinics or in clinical trials target diseases other than bacterial infections. Although several groups and companies are working on different strategies, the application of antisense technologies to prokaryotes still lags with respect to those that target other human diseases. In those cases where the focus is on bacterial pathogens, a subset of the research is dedicated to produce antisense compounds that silence or reduce expression of antibiotic resistance genes. Therefore, these compounds will be adjuvants administered with the antibiotic to which they reduce resistance levels. A varied group of oligonucleotide analogs like phosphorothioate or phosphorodiamidate morpholino residues, as well as peptide nucleic acids, locked nucleic acids and bridge nucleic acids, the latter two in gapmer configuration, have been utilized to reduce resistance levels. The major mechanisms of inhibition include eliciting cleavage of the target mRNA by the host’s RNase H or RNase P, and steric hindrance. The different approaches targeted resistance to β-lactams including carbapenems, aminoglycosides, chloramphenicol, macrolides, and fluoroquinolones.

An experiment was conducted to investigate the role of single nucleotide polymorphisms of the gene RNF34 3′UTR-588 G>A and RNF128 I1-2380C>T with carcass and meat quality traits of Chinese Simmental-cross steers. METHOD: We performed sequencing and restriction enzyme digestion to detect genotypes of RNF34 3′UTR-588 G>A and RNF128 I1-2380C>T. Then, we analyzed the association of novel single nucleotide polymorphisms in intron regions of the RNF128 gene and in the 3′UTR region of RNF34 and meat quality traits of Chinese Simmental-cross steers. RESULTS: Statistical analyses revealed that SNP of RNF128 (I1-2380C>T) was significantly associated with dressed weight, forepaw weight, carcass depth, carcass brisket depth, hind legs length (P<0.05), etc. And RNF34 (3′UTR-588 G>A) were significantly associated with testis weight, kidney weight, tare weight (P<0.05), etc. CONCLUSION: Our findings suggest that polymorphisms in RNF34 and RNF128 might be important genetic factors that influence carcass and meat quality in beef cattle. Thus, they might be useful markers for meat quality traits in future marker-assisted selection programs in beef cattle breeding and production.

We remind about the dogma initially established with the nucleic acid double helix, i.e. the DNA structure as the primary source of life. However, we bring into the discussion those additional processes that were crucial to enable life and cell evolution. Studying chemosensory proteins (CSPs) and odor binding proteins (OBPs) of insects, we have found a high level of pinpoint mutations on the RNA and peptide sequences. Many of these mutations are found to be tissue-specific and induce subtle changes in the protein structure, leading to a new theory of cell multifunction and life evolution. Here, attention is given to RNA and peptide mutations in small soluble protein families known for carrying lipids and fatty acids as fuel for moth cells. A new phylogenetic analysis of mutations is presented and provides even more support to the pioneer work, i.e. the finding that mutations in binding proteins have spread through moths and various groups of insects. Then, focus is given to specific mechanisms of mutations that are not random, change α-helical profilings and bring new functions at the protein level. In conclusion, RNA and peptide mutations are not seen as representative of a multitude of diseases, but rather as an alternative way by which protocells developed to acquire multifunction and totipotency. This provides a basis for the theory of RNA/peptide mutations for birth and evolution of life on earth’s crust proposed here.

We present simple relations for nuclear stability and nuclear binding energy with respect to three gravitational constants associated with electroweak, strong and electromagnetic interactions.

In this paper we propose the generation of synthetic small and more sophisticated molecule structures that optimize the binding affinity to a target (ASYNT-GAN). To achieve this we leverage on three important achievements in A.I.: Attention, Deep Learning on Graphs and Generative Adversarial Networks. Similar to text generation based on parts of text we are able to generate a molecule architecture based on an existing target. By adopting this approach, we propose a novel way of searching for existing compounds that are suitable candidates. Similar to question and answer Natural Language solutions we are able to find drugs with highest relevance to a target. We are able to identify substructures of the molecular structure that are the most suitable for binding. In addition, we are proposing a novel way of generating the molecule in 3D space in such a way that the binding is optimized. We show that we are able to generate compound structures and protein structures that are optimised for binding to a target.

The vast majority of patients with pancreatic ductal adenocarcinoma (PDAC) suffer cachexia. Although cachexia results from concurrent loss of adipose and muscle tissue, most studies focus on muscle alone. Emerging data demonstrate the prognostic value of fat loss in cachexia. Here we sought to identify the muscle and adipose gene profiles and pathways regulated in cachexia. Matched rectus abdominis muscle and subcutaneous adipose tissue were obtained at surgery from patients with benign conditions (n=11) and patients with PDAC (n=24). Self-reported weight loss and body composition measurements defined cachexia status. Gene profiling was done using Ion proton sequencing. Results were queried against external datasets for validation. 961 DE genes were identified from muscle and 2000 from adipose tissue, demonstrating greater response of adipose than muscle. In addition to known cachexia genes such as FOXO1, novel genes from muscle, including PPP1R8 and AEN correlated with cancer weight loss. All the adipose correlated genes including SCGN and EDR17 are novel for PDAC cachexia. Pathway analysis demonstrated shared pathways but largely non-overlapping genes in both tissues. Age related muscle loss predominantly had a distinct gene profiles compared to cachexia. This analysis of matched, externally validate gene expression points to novel targets in cachexia.

One of creators of quantum mechanics P. Jordan in his work on quantum biology claimed that life's missing laws were the rules of chance and probability of the quantum world. The article presents author’s results of studying frequencies (or probabilities) of nucleotides on so-called epi-chains of long DNA sequences of various eukaryotic and prokaryotic genomes. DNA epi-chains are algorithmically constructed subsequencies of DNA nucleotide sequences. According to the algorithm of construction of any epi-chain of the order n, the epi-chain is such nucleotide subsequence, in which the numerations of adjacent nucleotides differ by natural number n (n = 1, 2, 3, 4,…). Correspondingly each epi-chain of order n ≥ 2 contains n times less nucleotides than the original DNA sequence. The presented results unexpectedly discover that in long single-stranded and double-stranded DNA of any tested genome its DNA epi-chains of different orders n (values n are not too large) have practically identical frequencies (or probabilities) of each kind of nucleotides. These data allow considering DNA as a regular rich set of epi-chains, which can play a certain role in genetic and epigenetic phenomena as the author belives. Appropriate rules of nucleotide frequencies on epi-chains of long DNA sequences are formulated for further their tests on a wider set of genomes. These results testify on existence of long-range coherence in long DNA and remind the Fröhlich's theory of long-range coherence in biological systems. The phenomenological data are discussed from different standpoints: the DNA double helices and helical antennas with circular polarizations of electromagnetic waves; relations with the Fröhlich's theory; numerical analysis of DNA epi-chains under binary representations of nucleotides. Results are useful for developing quantum and algebraic biology.

Rett syndrome (RTT), a neurodevelopmental disorder, is mainly caused by mutations in methyl CpG-binding protein 2 (MECP2), which alter the functions of domains to either bind to methylated DNA or interact with a transcriptional co-repressor complex. It has been established that alterations in cyclin-dependent kinase-like 5 (CDKL5) or forkhead box protein G1 (FOXG1) correspond to distinct neurodevelopmental disorders, given that a series of studies have indicated that RTT is also caused by alterations in either one of these genes. We tried to elucidate RTT through evolution and structure assessment of MeCP2, CDKL5, and FOXG1, by focusing on their binding partners and disordered structures. Here, we provide insight into the similarities of the FOXG1 and MECP2 binding partners evolution and function. On the other hand, we suggest that CDKL5 could be a potential candidate for a classical RTT treatment, particularly based on its disordered structure that spans after the catalytic domain to the C-terminus, which shows abundant linear motifs that can bind to molecules with divergent structures of similar affinity. Additionally, we provide insight into the relationship between disordered structure and disease.

With reference to our earlier published views on large nuclear gravitational constant , nuclear elementary charge and strong coupling constant , in this paper, we present simple relations for nuclear stability range, binding energy of isotopes and magic proton numbers.

Tumor invasion, metastasis, and recrudesce remain a considerable challenge in the treatment of gastric cancer (GC). Herein, we first identified that RBFOX3 (RNA binding protein fox-1 homolog 3) was significantly up-regulated in GC tissues and negatively linked to the survival rate of GC patients. RBFOX3 promoted cell division and cell cycle progression in vitro as well as in vivo. Furthermore, RBFOX3 increased cell invasion and migration ability. Interestingly, both the suppression of GC cell multiplication and invasion moderated by the silencing of RBFOX3 was rescued by HTERT up-regulation. Additionally, RBFOX3 augmented the resistance of GC cells to 5-fluorouracil (5-Fu) by repressing RBFOX3. Mechanistically, exogenous up-regulation of RBFOX3 triggered promoter activity and HTERT expression thereby enhancing the division and development of GC cells. Importantly, our findings revealed that RBFOX3 interacted with AP-2β to modulate the HTERT expression as demonstrated by co-immunoprecipitation analysis. In conclusion, our study indicates that high expression of RBFOX3 promotes GC progression and development but predicts worse prognosis by stimulating HTERT signaling. Moreover, the results suggest that the RBFOX3/AP-2β/HTERT pathway is a novel target for the development of therapeutic agents for the prevention and treatment of GC reappearance and metastasis.

The selectivity in selective autophagy pathways is achieved via the selective autophagy receptors (SARs) – proteins that bind a ligand on the substrate to be degraded and the Atg8-family protein on the growing autophagic membrane, phagophore, effectively bridging them. In mammals, the most common ligand of SARs is ubiquitin, a small protein modifier that tags substrates for their preferential degradation by autophagy. Consequently, the most common SARs are the ubiquitin-binding SARs, such as SQSTM1/p62 (sequestosome 1). Surprisingly, there is only one SAR of this type in yeast – Cue5, which acts as a receptor for aggrephagy and proteaphagy – pathways that remove the ubiquitinated protein aggregates and proteasomes, respectively. However, recent studies described the ubiquitin-dependent autophagic pathways that do not require Cue5, e.g. stationary phase lipophagy for intracellular lipid droplets and nitrogen starvation-induced mitophagy for mitochondria. What is the role of ubiquitin in these pathways? Here, we propose that the ubiquitinated lipid droplets and mitochondria are recognized by the alternative ubiquitin-binding SARs. Our analysis identifies the proteins that could potentially fulfill this role in yeast. We think that matching of the ubiquitin-dependent (but Cue5-independent) autophagic pathways with the ubiquitin-and-Atg8-binding proteins enlisted here might uncover the novel ubiquitin-binding SARs in yeast.

Background: Recent developments in our understanding of the interactions between long non-coding RNA (lncRNA) and cellular components have improved treatment approaches for various human diseases including cancer, vascular diseases, and neurological diseases. Although investigation of specific lncRNAs revealed their role in the metabolism of cellular RNA, our understanding of their contribution to post-transcriptional regulation is relatively limited. In this study, we explore the role of lncRNAs in modulating alternative splicing and their impact on downstream protein-RNA interaction networks. Results: Analysis of alternative splicing events across 39 lncRNA wildtype and knockout RNA-sequencing datasets from three human cell lines: HeLa (Cervical Cancer), K562 (Myeloid Leukemia), and U87 (Glioblastoma), resulted in high confidence (fdr < 0.01) identification of 4432 skipped exon events and 2474 retained intron events, implicating 759 genes to be impacted at post-transcriptional level due to the loss of lncRNAs. We observed that a majority of the alternatively spliced genes in a lncRNA knockout were specific to the cell type, in agreement with the finding that genes affected by alternative splicing also displayed enriched functions in a cell type specific manner. To understand the mechanism behind this cell-type specific alternative splicing patterns, we analyzed RNA binding protein (RBP)-RNA interaction profiles across the spliced regions. Conclusions: Despite limited RBP binding data across cell lines, alternatively spliced events detected in lncRNA perturbation experiments were associated with RBPs binding in proximal intron-exon junctions, in a cell type specific manner. The cellular functions affected by alternative splicing were also affected in a cell type specific manner. Based on the RBP binding profiles in HeLa and K562 cells, we hypothesize that several lncRNAs are likely to exhibit a sponge effect in disease contexts, resulting in the functional disruption of RBPs due to altered titration of the RBPs from their target loci. We propose that such lncRNA sponges can extensively rewire the post-transcriptional gene regulatory networks by altering the protein-RNA interaction landscape in a cell-type specific manner.

Two X chromosomes of female mammals randomly inactivate one of paternal or maternal X chromosome in early embryonic development and all the daughter cells produced from these cells retain the same feature of X chromosome inactivation, which is called X chromosome inactivation (XCI). Studying the mechanisms of XCI is important for understanding epigenetic that plays an important role in age-associated diseases. The previous studies have demonstrated that binding of RNAs and DNAs may play a role in activating gene expression. In this paper, our study aims to explore whether the mechanisms of XCI involve the RNA binding strength to X chromosome DNAs. The bioinformatics analyses based on big data were used to analyze the simulated binding strength of RNAs (RNA binding strength) to 23 chromosomes (including X chromosome and 22 human autosomes) and the characteristics of repetitive sequences in the X-inactivation centre. The results revealed that RNA binding strength of the long arm of the X chromosome that is almost entirely inactivated in XCI was significantly lower than that of all autosomes and the short arm of X chromosome, meanwhile the RNA binding strengths of inactivation regions in X chromosome were significantly lower than that of regions escaping from XCI. Different repetitive sequence clusters within the center of XCI presented a cross distribution characteristic. To further prove whether the repetitive sequences in human X chromosome involve in XCI, we cloned long interspersed element (LINE-1, L1) and short interspersed element (Alu) from human Xq13, the center of XCI, and constructed expression vectors carrying sense-antisense combination repetitive sequences (L1s or Alus). Effects of combined L1 or combined Alu sequences on expression of EGFP reporter gene were examined in stably transfected HeLa cells, which simulates the effects of repetitive sequences located on chromosomes. The results of experiments revealed transcribed L1 repetitive sequences activated EGFP reporter gene expression, so did the Alus. The experiment results suggested repetitive sequences activated genes by interaction of transcribed RNAs and DNAs. Since the binding of RNAs and DNAs can activate gene, so the low RNA binding strength of human X chromosome may be one of reasons of XCI. The cross distribution characteristics of different repetitive sequence clusters leading to a cascade of gene activation or gene inactivation may be the reason of transcriptional silencing one of the X chromosomes in female mammals.

Amyotrophic Lateral Sclerosis (ALS) is an irreversible neurodegenerative disease with no known cure. Recent studies suggest a strong metabolic component in ALS pathogenesis and have shown an inverse relationship between leptin levels and ALS progression, although the effects of leptin as a treatment have not yet been studied. Therefore, we aim to examine whether the acute treatment with leptin has beneficial effects on brain pathology and cognitive function in the transgenic TDP43A315T line of ALS. Mice were treated intranasally (IN) with 0.03mg/kg of leptin or vehicle (VH) daily for 7 days. Data showed a progressive decline in body weight and motor coordination in TDP43A315T mice. Moreover, Lep-treated TDP43A315T mice showed an earlier disease onset, along with an improvement in motor performance. Altered levels of some of the adipokines and metabolic proteins studied were found in TDP43A315T mice, which were differently expressed among Lep-treated and VH-treated animals. Furthermore, some correlations were found among the serum levels of this proteins in WT and TDP43A315T mice. As far as we know, this is the first pilot study to provide evidence of the therapeutic effect of leptin treatment in a mice model of ALS, although further studies are needed to expound on the underlying mechanisms.

Background: Cytokines are multifunctional glycoproteins playing a vital role in the tumor microenvironment and in the progression of breast cancer. The immune responses in the tumors restrained by pro- and anti-inflammatory cytokine expressions could be influenced by genetic polymorphisms. Hence, the present study evaluated the contribution of IL6 (rs1800797, rs1800796, and rs1800795), and IL18 (rs1946518, rs187238, and rs549908) genotypes and its haplotypes with risk and progression of breast cancer in south Indian population. Methods: The polymorphisms of IL6 gene -597G>A, -572G>C, -174G>C and IL18 gene -607C>A, -137G>C, and 105A>T were genotyped through PCR-RFLP and As-PCR assays in blood DNA of 600 subjects. We have performed haplotype, LD, univariate, multivariate logistic regression and Kaplan-Meier analyses for the obtained data. Results: The frequency of AA genotype & A-allele of IL6 -597G>A, and CC genotype & C allele of IL6 -174G>C polymorphism was higher in breast cancer patients and was found to be significantly associated with TNM staging, late (advanced) stage, metastasis, etc. While, CG and GG genotypes of IL6 -572 C>G polymorphism had a protective role against breast cancer. Further, IL18 -607C>A, -137G>C & 105A>T polymorphisms were found to be associated with lobular carcinoma subtype, PR-ve and HER2+ve breast cancer patients. Perfect LD was observed between all SNPs of IL6 & IL18 genes under study; G-C-C, A-G-G and A-C-C haplotype combination of IL6 genes had conferred 2.09, 2.25 and 4.72 folds risk for breast cancer respectively. In survival analysis, we observed that the C allele of rs 1800795 was found to be significantly associated with 5years overall survival in breast cancer subjects. Conclusions: Overall, our results suggest the importance of genotypic and haplotype analysis of IL6, and IL18 gene variants in progression and risk identification of breast cancer.

Substrate binding proteins (SBP) bind to specific ligands in the periplasmic region and bind to membrane proteins to participate in transport or signal transduction. Typical SBPs consist of two α/β domains and recognize the substrate by hinge motion between two domains. Conversely, short length Rhodothermus marinus SBP (named as RmSBP) exists around the methyl-accepting chemotaxis protein. We previously determined the crystal structure of RmSBP consisting of a single α/β domain, but the substrate recognition mechanism is still unclear. To better understand the short length RmSBP, we performed comparative structure analysis, computational substrate docking, and X-ray crystallographic study. RmSBP shares a high level of similarity in α/β domain with other SBP proteins, but it has a distinct topology in the C-term region. The substrate binding model suggested that conformational change in the peripheral region of RmSBP was required to recognize the substrate. We determined the crystal structures of RmSBP at pH 5.5, 6.0, and 7.5. RmSBP showed structural flexibility of the β1-α2 loop, β5-β6 loop, and extended C-term domain based on the electron density map and temperature B-factor analysis. These results provide information that will further the understanding of the function of the short length SBP.

Pteridine reductase 1 is a trypanosomatid multifunctional enzyme that provides a mechanism for escape of Dihydrofolate reductase (DHFR) inhibition. This is because PTR1 can reduce pterins and folates. Trypanosomes require folates and pterins for survival and are unable to synthesize them de novo. Currently there are no anti-folate based Human African Trypanosomiasis (HAT) chemotherapeutics in use. Thus, successful dual inhibition of TbDHFR and TbPTR1 has implications in the exploitation of anti-folates. We carried out molecular docking of a ligand library of 5742 compounds against TbPTR1 and identified 18 compounds showing promising binding modes. The protein-ligand complexes were subjected to Molecular dynamics to characterize their molecular interactions and energetics followed by in vitro testing. In this study, we identified five potential TbPTR1 inhibitors that showed low micromolar Trypanosome growth inhibition in in vitro experiments with no significant human cell cytotoxicity. Compounds RUBi004, RUBi007, RUBi014, and RUBi018 displayed moderate to strong antagonism when used in combination with the known TbDHFR inhibitor, WR99210. This gave an indication that the compounds might inhibit both TbPTR1 and TbDHFR. RUBi016 showed an additive effect in the isobologram assay. Our results provide a basis for scaffold optimization for further studies in the development of HAT antifolates.

Earlier studies reported alterations of the kynurenine (KYN) pathway of tryptophan (TRP) metabolism in Parkinson’s disease (PD). The first rate-limiting enzymes indoleamine 2,3-dioxygenase (IDO) and tryptophan dioxygenase were observed upregulated, resulting elevated KYN/TRP ratios in the serum and cerebrospinal fluid samples of patients with PD. An increasing number of single nucleotide polymorphisms (SNPs) have been identified in a population of PD. However, little is known if genetic variations of the IDO contribute to disturbance of the KYN metabolism in and the pathogenesis of PD. SNP analysis of IDO1 was performed by allelic discrimination assay with fluorescently labelled TaqMan probes and a subgroup analysis was conducted according to the age of PD onset. The frame shifts variant rs34155785, intronic variant rs7820268, and promotor region variant rs9657182 SNPs of 105 PD patients without comorbidity were analyzed and compared to 129 healthy controls. No significant correlation was found in three SNPs between PD patients and healthy controls. However, the subgroup analysis revealed that A alleles of rs7820268 SNP or rs9657182 SNP carriers contribute to later onset of PD than non-carriers. The study suggested that SNPs of IDO1 influenced the age onset of PD and genotyping of SNPs in certain alleles potentially serves as a risk biomarker of PD.

The recognition of (1→3)-β-D-glucans (BGs) by β-1,3-D-glucan recognition protein (BGRP) found in invertebrates plays a significant role in the activation of toll pathway and pro-phenol oxidase system in insect host defense against fungal invasion. To examine the structural diversity of BGs in BGRP interaction, the binding specificity of BGRPs cloned from four different insectswas characterized using ELISA. Recombinant BGRPs expressed as Fc-fusion proteins of human IgG1 bound to solid phase BGs. Because of the binding specificities, the BGRPs were categorized into two different ultrastructure- binding characters. The BGRPs from Silkworm and Indian meal moth bound to BGs containing triple-helical structure. Other BGRPs from red flour beetle and yellow mealworm beetle showed no binding to triple-helical BGs, but to alkaline-treated BGs, which have partially opened helical conformation. These evidences suggest that the innate immune system distinguishes different BG conformations and it is equipped for the diversity of BG structures.

Detection of PLA2R and THSD7A among primary membranous glomerulonephritis (MGN) patients transformed the diagnosis, treatment monitoring and prognosis. Anti-PLA2R can be detected in 70-90% of primary MGN patients while anti-THSD7A in 2-3% of anti-PLA2R negative primary MGN patients depending on the technique used. Serum and urine samples are less invasive and non-invasive respectively and can detect the presence of anti-PLA2R and anti-THSD7A with higher sensitivity and specificity, significant in patients’ monitoring and prognosis better than exposing patients to frequent biopsy which is an invasive procedure. Different techniques of detection of PLA2R and THSD7A in patients’ urine and sera were reviewed with the aim of providing newer and alternative techniques. We proposed the use of biomarkers (PLA2R and THSD7A) in making the diagnosis, treatment decision and follow up of patients with primary MGN. We also reviewed other prognostic renal biomarkers like retinol binding protein (RBP) and beta-2 microglobulin in order to detect progression of renal damage for early intervention.

Mitochondrial dysfunction is relevant mechanism in cardiac aging. Here, we investigated the effects of late-life enalapril administration at non-antihypertensive dose on mitochondrial genomic stability, oxidative damage, and mitochondrial quality control (MQC) signaling in the heart of aged rats. The protein expression of selected mediators (i.e., mitochondrial antioxidant enzymes, energy metabolism, mitochondrial biogenesis, dynamics, and autophagy) was measured in old rats randomly assigned to receive enalapril (n=8) or placebo (n=8) from 24 to 27 months of age. We also assessed mitochondrial DNA (mtDNA) content, citrate synthase activity, oxidative lesions to protein and mtDNA (i.e., carbonyls and abundance of mtDNA4834 deletion), and mitochondrial transcription factor A (TFAM) binding to specific mtDNA regions. Enalapril attenuated cardiac hypertrophy and oxidative stress-derived damage (mtDNA oxidation, mtDNA4834 deletion, and protein carbonylation), while increasing mitochondrial antioxidant defenses. TFAM binding to mtDNA regions involved in replication and deletion generation was increased following enalapril administration. Increased mitochondrial mass as well as mitochondriogenesis and autophagy signaling was found in enalapril-treated rats. Late-life enalapril administration mitigates age-dependent cardiac hypertrophy and oxidative damage, while increasing mitochondrial mass and modulating MQC signaling. Further analyses are needed to conclusively establish whether enalapril may offer cardioprotection during aging.

Pathogenicity is unknown for the majority of human gene variants. For prioritization of sequenced somatic and germline mutation variants, in silico approaches can be utilized. In this study, 84 million non-synonymous Single Nucleotide Variants (SNVs) in the human coding genome were annotated using consensus Variant Effect Prediction (cVEP) method. An algorithm, implemented as a stacked ensemble of supervised learners, performed combination of the 39 functional, conservation mutation impact scores from dbNSFP4.0. Adding gene indispensability score, accounting for differences in the pathogenicities of the variants in the essential and the mutation-tolerant genes, improved the predictions. For each SNV the consensus combination gives either a continuous-value pathogenicity score, or a categorical score in five classes: pathogenic, likely pathogenic, uncertain significance, likely benign, benign. The provided class database is aimed for direct use in clinical practice. The trained prediction models were 5-fold cross-validated on the evidence-based categorical annotations from the ClinVar database. The rankings of the scores based on their ability to predict pathogenicity were obtained. A two-step strategy using the rankings, scores and class annotations is suggested for filtering and prioritization of the human exome mutations in clinical and biological applications of NGS technology.

Over 250 million people are infected chronically with hepatitis B virus (HBV), the leading cause of liver cancer worldwide. HBV persists due in part to its compact, stable minichromosome, the covalently-closed, circular DNA (cccDNA), which resides in the hepatocytes’ nuclei. Current therapies target downstream replication products, however, a true virological cure will require targeting the cccDNA. Finding targets on such a small, compact genome is challenging. For HBV, to remain replication-competent, it needs to maintain nucleotide fidelity in key regions, such as the promoter regions, to ensure that it can continue to utilize the necessary host proteins. HBVdb (HBV database) is a repository of HBV sequences spanning all genotypes (A-H) amplified from clinical samples, and hence implying an extensive collection of replication-competent viruses. Here, we analyzed the HBV sequences from HBVdb using bioinformatics tools to comprehensively assess the HBV core and X promoter regions amongst the nearly 70,000 HBV sequences for highly-conserved nucleotides and variant frequencies. Notably, there is a high degree of nucleotide conservation within specific segments of these promoter regions highlighting their importance in potential host protein-viral interactions and thus the virus’ viability. Such findings may have key implications for designing antivirals to target these areas.

Diarrhoea caused by enterotoxigenic Escherichia coli is one of the leading causes of mortality in children under five years of age and is a great burden on developing countries. The major virulence factor of the bacterium is the heat-labile enterotoxin (LT), a close homologue of the cholera toxin. The toxins bind to carbohydrate receptors in the gastrointestinal tract, leading to toxin uptake and, ultimately, to severe diarrhoea. Previously, LT from human- and porcine-infecting ETEC (hLT and pLT, respectively) were shown to have different carbohydrate-binding specificities, in particular with respect to N-acetyllactosamine-terminating glycosphingolipids. Here, we probed eleven single-residue variants of the heat-labile enterotoxin with surface plasmon resonance spectroscopy and compared the data to the parent toxins. In addition we present a 1.45 Å crystal structure of pLTB in complex with branched Lacto-N-neohexaose (Galbeta4GlcNAcbeta6[Galbeta4GlcNAcbeta3]Galbeta4Glc). The largest difference in binding specificity is caused by mutation of residue 94, which links the primary and secondary binding sites of the toxins. Residue 95 (and to a smaller extent also residues 7 and 18) also contribute, whereas residue 4 shows no effect on monovalent binding of the ligand and may rather be important for multivalent binding and avidity.

A new label-free molecular probe for luminescent nucleotide detection in neutral aqueous solution is presented. Phosphate-containing molecules such as nucleotides possess vital role in cell metabolism, energy economy, and signaling. Thus the monitoring of nucleotide concentration and nucleotide related enzymatic reactions is of high importance. Two component lanthanide complex formed from Tb(III) ion carrier and light harvesting antenna, readily distinguishes nucleotides containing different number of phosphates and enable direct detection of enzymatic reactions converting nucleotide triphosphate (NTP) to nucleotide di/monophosphate or the opposite. Developed sensor enables the detection of enzymatic activity with a low nanomolar sensitivity, as highlighted with K-Ras and apyrase enzymes in there hydrolysis assays performed in high throughput screening compatible 384-well plate format.

Cells form a bipolar spindle during mitosis to ensure accurate chromosome segregation. Proper spindle architecture is established by a set of kinesin motors and microtubule-associated proteins. In most eukaryotes, kinesin-5 motors are essential for this process, and genetic or chemical inhibition of their activity leads to the emergence of monopolar spindles and cell death. However, these deficiencies can be rescued by simultaneous inactivation of kinesin-14 motors, as they counteract kinesin-5. We conducted detailed genetic analyses in fission yeast to understand the mechanisms driving spindle assembly in the absence of kinesin-5. Here we show that deletion of the nrp1 gene, which encodes a putative RNA-binding protein with unknown function, can rescue temperature sensitivity caused by cut7-22, a fission yeast kinesin-5 mutant. Interestingly, kinesin-14/Klp2 levels on the spindles in the cut7 mutants were significantly reduced by the nrp1 deletion, although the total levels of Klp2 and the stability of spindle microtubules remained unaffected. Moreover, RNA-binding motifs of Nrp1 are essential for its cytoplasmic localization and function. We have also found that a portion of Nrp1 is spatially and functionally sequestered by chaperone-based protein aggregates upon mild heat stress and limits cell division at high temperatures. We propose that Nrp1 might be involved in post-transcriptional regulation through its RNA-binding ability to promote the loading of Klp2 on the spindle microtubules.

The worldwide outbreak of a novel coronavirus, SARS-CoV-2 has caused a pandemic of respiratory disease. Due to this emergency, researchers around the globe have been investigating the evolution of the genome of SARS-CoV-2 in order to design vaccines. Here I examined the evolution of GC content of SARS-CoV-2 by comparing the genomes of the members of the group Betacoronavirus. The results of this investigation revealed a highly significant positive correlation between the GC contents of betacoronaviruses and their divergence from SARS-CoV-2. The betacoronaviruses that are distantly related to SARS-CoV-2 have much higher GC contents than the latter. Conversely, the closely related ones have low GC contents, which are only slightly higher than that of SARS-CoV-2. This suggests a systematic reduction in the GC content in the SARS-CoV-2 lineage over time. The declining trend in this lineage predicts a much-reduced GC content in the coronaviruses that will descend/evolve from SARS-CoV-2 in the future. Due to the three consecutive outbreaks (MERS-CoV, SARS-CoV and SARS-CoV-2) caused by the members of the SARS-CoV-2, the scientific community is emphasizing the need for universal vaccines that are effective across many strains including those, that will inevitably emerge in the near future. The reduction in GC contents implies an increase in the rate of GC→AT mutations than that the mutational changes in the reverse direction. Therefore, understanding the evolution of base composition and mutational patterns of SARS-CoV-2 could be useful in designing broad-spectrum vaccines that could identify and neutralize the present and future strains of this virus.

The accurate virus detection, strain discrimination, and source attribution of contaminated food items remains a persistent challenge because of the high mutation rates anticipated to occur in foodborne RNA viruses, such as Hepatitis A virus (HAV). This has led to predictions of the existence of more than one sequence variant between the hosts (inter-host) or within an individual host (intra-host). However, there have been no reports of intra-host variants from an infected single individual, and little is known about the accuracy of the single nucleotide variations (SNVs) calling with various methods. In this study, the presence and identity of viral SNVs, either between HAV clinical specimens or among a series of samples derived from HAV clone1-infected FRhK4 cells, were determined following analyses of nucleotide sequences generated using next-generation sequencing (NGS) and pyrosequencing methods. The results demonstrate the co-existence of inter- and intra-host variants both in the clinical specimens and the cultured samples. The discovery and confirmation of multi-viral RNAs in an infected individual is dependent on the strain discrimination at the SNV level, and critical for successful outbreak traceback and source attribution investigations. The detection of SNVs in a time series of HAV infected FRhK4 cells improved our understanding on the mutation dynamics determined probably by different selective pressures. Additionally, it demonstrated that NGS could potentially provide a valuable investigative approach toward SNV detection and identification for other RNA viruses.

Adipocytes play a critical role in maintaining a healthy systemic metabolism by storing and releasing energy in the form of fat and helping to regulate glucose and lipid levels in the body. Adipogenesis is the process through which pre-adipocytes are differentiated into mature adipocytes. It is a complex process involving various transcription factors and signaling pathways. Dysregulation of adipogenesis has been implicated in the development of obesity and metabolic disorders. Therefore, understanding the mechanisms that regulate adipogenesis and the factors that contribute to its dysregulation may provide insights into the prevention and treatment of these conditions. RNA Binding Motif Single Stranded Interacting Protein 1 (RBMS1) is a protein that binds to RNA and plays a critical role in various cellular processes such as alternative splicing, mRNA stability, and translation. The RBMS1 polymorphism has been shown to be associated with obesity and Type 2 diabetes, but the role of RBMS1 in adipose metabolism and adipogenesis is not known. We show that RBMS1 is highly expressed during the early phase of differentiation of the murine adipocyte cell line 3T3-L1 and is significantly upregulated in adipose tissue depots and adipocytes of high-fat-fed mice, implying a possible role in adipogenesis and adipose metabolism. Knockdown of RBMS1 in pre-adipocytes impacted the differentiation process and reduced the expression of some of the key adipogenic markers. Transcriptomic and proteomic analysis indicated that RBMS1 depletion affected the expression of several genes involved in major metabolic processes, including carbohydrate and lipid metabolism. Our findings imply that RBMS1 plays an important role in adipocyte metabolism and may offer novel therapeutic opportunity for metabolic disorders such as obesity and type 2 diabetes.

Since the detection of the first COVID-19 patient, 2 years have passed, during which more than 287,862,000 people fell ill globally, of which about 1.9% died. Implementation of SARS-CoV-2 control programs required efforts from almost all countries. An important direction in the fight against COVID-19 was the formation of herd immunity, the main tool for managing the pandemic. Study goal: to assess the seroprevalence of antibodies (Abs) to SARS-CoV-2 nucleocapsid (Nc) and receptor binding domain (RBD) in the St. Petersburg population during the COVID-19 pandemic. Materials and methods. A longitudinal cohort randomized monitoring study of Ab seroprevalence (SARS-CoV-2 Nc, RBD) was organized and conducted according to a unified methodology developed by Rospotrebnadzor with the participation of the St. Petersburg Pasteur Institute. For this purpose, a cohort of 1000 volunteers was formed who participated in all five stages of seromonitoring. The cohort was divided into 7 age groups: 1-17; 18-29; 30-39; 40-49; 50-59; 60-69; 70; and older (70+) years. Seropositivity levels (Nc, RBD) were assessed by quantitative and qualitative enzyme immunoassays. During the 2nd year of monitoring, some volunteers were vaccinated with the GamCOVIDVac (84%) or EpiVacCorona (11.6%) vaccines approved in Russia. Statistical processing was carried out using the Excel 2010 software package. Confidence intervals for shares and percentages (95% CI) were calculated using the method of A. Wald and J. Wolfowitz with adjustment (A. Agresti, B.A. Coull). The statistical significance of differences was calculated by z-test, using the appropriate online calculator (p<0.05), unless indicated. Results. There was a trend towards: an increase in Nc seropositivity in stages 1-3 of seromonitoring, with a decrease in stages 4-5 among children and adults. The share of RBD seropositive steadily increased during all 5 stages of seromonitoring. The most frequently found were low anti-RBD Abs levels (22.6-220 BAU/ml). High Ab levels were recorded statistically significantly less frequently. Asymptomatic forms were observed in 84-88% of SARS-CoV-2 seropositive volunteers. By the 5th stage of monitoring, this indicator significantly decreased to 69.8% (95% CI: 66.1-73.4). The monitoring revealed a statistically significant increase in anti-RBD Abs, alongside a statistically significant decrease in the proportion of Nc seropositive. This dynamic was especially characteristic of persons vaccinated with GamCOVIDVac. Conclusion. Prior to the use of specific vaccines, a seroprevalence of anti-Nc Abs was noted. After the introduction of the GamCOVIDVac vaccine in adults, a decrease in the level of anti-Nc Abs was noted due to an increase in the proportion of RBD seropositive persons.

Demonstrating biosimilarity entails comprehensive analytical evaluations, clinical pharmacolo-gy profiling, and efficacy testing for at least one medical indication in patients. These require-ments are stipulated by the U.S. Biologics Price Competition and Innovation Act (BPCIA). The costliest element—efficacy testing—can be waived if other compliance benchmarks are satisfied, including comparing functional pharmacodynamic (PD) biomarkers, even when they do not di-rectly correlate with clinical outcomes. Most biological drugs, such as monoclonal antibodies (mAbs), lack identifiable PD biomarkers. The FDA has employed various 'omics' technologies to identify potential PD biomarkers, including proteomics, glycomics, transcriptomics, genomics, epigenomics, and metabolomics. Although these efforts provide a robust scientific basis for estab-lishing biosimilarity, they are neither practical nor necessarily superior to existing functional biomarkers, such as receptor binding and mode-of-action outcomes. As we report for the first time, these functional biomarkers can effectively serve as PD indicators for all FDA-licensed bio-logical drugs. We recommend that the FDA consider officially listing these functional biomarkers to expedite and reduce the cost of biosimilar development, thereby increasing the accessibility of biological drugs. PD surrogates, like the receptor binding and pharmacokinetic profiles, are more robust and offer a rational solution to finding PD markers to compare for establishing biosimi-larity.

Enterovirus 71 (EV71) infection hand-foot-mouth disease (HFMD), meningoencephalitis, neonatal sepsis, and even fatal encephalitis in children, thereby representing a serious public health hazard. It is important to determine the mechanisms underlying the regulation of EV71 infection. In this study, we initially reveal that the interleukin enhancer binding factor 2 (ILF2) down-regulates EV71 50% tissue culture infective dose (TCID50), attenuates EV71 plaque formation unit (PFU), thereby repressing EV71 infection. Moreover, we reveal a distinct mechanism by which EV71 antagonizes ILF2-mediated antiviral effects. Chip data analyses show that ILF2 mRNA is reduced upon EV71 infection. Cellular studies indicate that EV71 infection represses ILF2 mRNA expression and protein production in human leukemic monocytes (THP-1) differentiated macrophages and in human rhabdomyosarcoma (RD) cells. Additionally, EV71 non-structural protein 2B interacts with ILF2 in human embryonic kidney (HEK293T) cells. Interestingly, in the presence of EV71 2B, ILF2 is translocated from the nucleus to the cytoplasm and co-localizes with 2B in the cytoplasm. Therefore, we reveal a distinct mechanism by which EV71 antagonizes ILF2-mediated antiviral effects by inhibiting ILF2 expression and promoting ILF2 translocation from the nucleus to cytoplasm through its 2B protein.

Pasmo (Septoria linicola) is a fungal disease causing major losses in seed yield and quality, and stem fibre quality in flax. Pasmo resistance (PR) is quantitative and has low heritability. To improve PR breeding efficiency, the accuracy of genomic prediction (GP) was evaluated using a diverse worldwide core collection of 370 accessions. Four marker sets, including three defined by 500, 134, and 67 previously identified quantitative trait loci (QTL) and one of 52,347 PR-correlated genome-wide single nucleotide polymorphisms, were used to build ridge regression best linear unbiased prediction (RR-BLUP) models using pasmo severity (PS) data collected from field experiments performed during five consecutive years. With five-fold random cross-validation, GP accuracy as high as 0.92 was obtained from the models using the 500 QTL when the average PS was used as the training dataset. GP accuracy increased with training population size, reaching values >0.9 with training population size greater than 185. Linear regression of the observed PS with the number of positive-effect QTL in accessions provided an alternative GP approach with an accuracy of 0.86. The results demonstrate the GP models based on marker information from all identified QTL and the 5-year PS average is highly effective for PR prediction.

To understand the mystery of final unification, in our earlier publications, we proposed two bold concepts: 1) There exist three atomic gravitational constants associated with electroweak, strong and electromagnetic interactions. 2) There exists a strong elementary charge in such a way that its squared ratio with normal elementary charge is close to reciprocal of the strong coupling constant. In this paper we propose that, can be considered as a compound physical constant associated with proton mass, electron mass and the three atomic gravitational constants. With these ideas, an attempt is made to understand nuclear stability and binding energy. In this new approach, nuclear binding energy can be fitted with four simple terms having one unique energy coefficient with a formula, where is an estimated mean stable mass number. With this new approach, Newtonian gravitational constant can be estimated in a verifiable approach with a model relation of the form, where is the Fine structure constant. Estimated and is 62 ppm higher than the CODATA recommended It needs further investigation. Proceeding further, an attempt is made to fit the recommended quark masses.

The mammalian/mechanistic target of rapamycin complex 1 (mTORC1) is activated by the small G-protein, RHEB-GTPase. On lysosome, RHEB activates mTORC1 by binding the domains of N-heat, M-heat, and FAT, which allosterically regulates ATP binding in the active site for further phosphorylation. The crucial role of RHEB in regulating growth and survival through mTORC1, makes it a targetable site for anti-cancer therapeutics. However, the binding kinetics of RHEB to mTORC1 is still unknown at the molecular level. Therefore, we studied the kinetics by in vitro and in-cell protein-protein interaction (PPI) assays. For this, we used the split-luciferase system (NanoBiT®) for in-cell studies, and prepared proteins for the in vitro measurements. Consequently, it was shown that RHEB binds to the whole mTOR both in the presence or absence of GTPɣS, with five-fold weaker affinity in the presence of GTPɣS. Also, RHEB bound to the truncated mTOR fragments of N-heat domain (60-167) and M-heat domain (967-1023) in a GTP independent manner. Furthermore, RHEB bound to the truncated kinase domain (2148-2300) with higher affinity also in GTP independently. In conclusion, RHEB binds two different binding sites of mTOR, which probably regulates the kinase activity of mTOR through multiple different molecular mechanisms.

RNA viruses exhibit a great tendency to mutate. Starting from 2019, the wild type that was labeled Hu-1 has during the last 3 years evolved to produce several dozen new variants, as a consequence of mutations. Mutations cause changes in empirical formulas of new virus strains, which lead to change in thermodynamic properties of biosynthesis and binding. These changes cause changes in the rate of reactions of binding of virus antigen to the host cell receptor and the rate of virus multiplication in the host cell. Changes in thermodynamic and kinetic parameters lead to changes in biological parameters of infectivity and pathogenicity. SARS-CoV-2 has starting from in 2019, until today, evolved towards increase in infectivity and maintaining constant pathogenicity, or for some variants a slight decrease in pathogenicity. In the case of Omicron BQ.1, BQ.1.1, XBB and XBB.1 variants pathogenicity is identical as in the Omicron BA.2.75 variant. On the other hand, infectivity of the Omicron BQ.1, BQ.1.1, XBB and XBB.1 variants is greater than those of previous variants. This will most likely result in the phenomenon of asymmetric coinfection, that is circulation of several variants in the population, some being dominant.

Little is known about the association between equol and bioavailable testosterone (BT) in adults. We examined the associations of urinary equol concentrations with serum total, bioavailable and free testosterone (FT), dehydroepiandrosterone sulfide (DHEAS), free androgen index (FAI) and sex hormone-binding globulin (SHBG) concentrations. This cross-sectional study included 1904 women aged 59.7 years. Urinary equol and serum sex hormone concentrations were measured. Overall, urinary equol tended to be inversely associated with bioactive forms of androgenic indices (BT, FT or FAI) but not with total testosterone (TT) or DHEAS. Urinary equol was also positively associated with SHBG. In multi-covariate-adjusted analyses stratified by menopausal status, graded and inverse associations between urinary equol and bioactive forms of androgenic indices (BT, FT and FAI) were observed in postmenopausal women (all p-trends <0.05), but not in premenopausal women. A significant positive association between urinary equol and SHBG was observed only in postmenopausal women. No significant associations were observed between urinary equol and TT or DHEAS in either group. A path analysis indicated that these associations of equol with androgens in postmenopausal women might be mediated by SHBG. Our findings indicated urinary equol exhibited graded and inverse associations with BT or FT but not TT in women.

Several animal food allergens assembly into amyloids under gastric-like environments. These aggregated structures provide Gad m 1 with an enhanced IgE interaction due to the amyloid assembly of the epitope regions. However, whether these properties are unique of Gad m 1 or common to other food allergens has not yet been addressed. Using Bos d 5, Bos d 12 and Gal d 2 as food allergen models and Gad m 1 as control, aggregation reactions and the sera of milk, egg and fish allergic patients we have analyzed the IgE interaction of the distinct amyloids. We found that amyloids formed by Bos d 12 and Gal d 2 full-length and truncated chains are recognized by the IgEs of milk and egg allergic patient sera. As with Gad m 1, in most cases amyloid recognition is higher than that of the precursor structure. Bos d 5 was not recognized under any fold by the IgE of the sera studied. These results support that formation of IgE-binding amyloids might be a common feature to animal food allergen.