Erlotinib inhibits epithelial growth factor receptor (EGFR) kinase activity and is used to treat non-small cell lung cancer (NSCLC). Despite its high efficacy, recurrence can occur in patients who become resistant to the drug. To address the underlying mechanism of Erlotinib resistance, we investigated additional mechanisms related to mode-of-drug-action, by multiple protein-binding interactions, besides EGFR by using drug affinity responsive target stability (DARTS) and liquid chromatography-mass spectrometry (LC-MS/MS) methods with non-labeled Erlotinib. DNA polymerase alpha subunit B (POLA2) was identified as a new Erlotinib binding protein that was validated by the DARTS platform, complemented with cellular thermal shift assays. Genetic knock-down of POLA2 promoted the anti-proliferative effect of the drug in the Erlotinib-resistant cell line H1299 with high POLA2 expression, whereas overexpression of POLA2 restored anti-proliferative effects in the Erlotinib-sensitive cell line HCC827 with low POLA2 expression. Importantly, POLA2 expression levels in four NSCLC cell lines were positively correlated with anti-proliferative Erlotinib efficacy, verified by bio-statistical analysis (Pearson correlation coefficient, R=0.9886). These results suggest that POLA2 is a novel complementary target protein of Erlotinib, and could clinically provide validity as a surrogate marker for drug resistance in patients with NSCLC.

A plenty of theories on the origin of genetic codes have been proposed so far, yet all ignored its energetic relation to the biochemical system. Here, a new hypothesis is proposed, according to which ATP is at the origin of the primordial genetic code by its coevolution with the pristine biochemical system. This hypothesis aims to show how the genetic code was produced by photochemical reactions in a protocell that derived from a lipid vescile enclosing various life’s building blocks (e.g. nucleotides and peptides). At extant cell, ATP is the only energetic product of photosynthesis, and is at the energetic heart of the biochemical systems. ATP could energetically elongate chains of both polynucleotides and polypeptides, thus providing a bridge between these bio-polymers and eventually mediating cellular biochemical innovation from energy transformation to informatization. ATP was not the only one that could drive the formation of polynucleotides and polypeptides, but favored by precellular selection. The protocell innovated a photosynthesis system to produce ATP efficiently and regularly with the aids of proteins and RNA/DNA. The completion of the genetic code from RNA to DNA marked the dawn of cellular life operated by Darwinian evolution. The ATP hypothesis supports the photochemical origin of life, shedding light on the formation of both photosynthetic and biochemical systems, which remains largely unknown thus far.

Purinergic receptors (P2X) are ATP-gated ion channels with an elusive role in the CNS. While the P2X2 and P2X4 subtypes are widely expressed in most neurons (i.e. at the edge of the postsynaptic densities of excitatory synapses), the direct contribution of P2X to synaptic transmission is uncertain. Several P2X have been shown to participate in receptor cross-talk: an interaction where one receptor (e.g. P2X2) influences the activity of another (e.g. GABA or AMPA receptors.) In this study, we tested for interactions between P2X2 or P2X4 and N-methyl-D-aspartate receptors (NMDARs). Using two-electrode voltage-clamp electrophysiology experiments in X. laevis oocytes, we demonstrate that both P2X2 and P2X4 interact with NMDARs in an inhibitory manner. When investigating the molecular domains responsible for this phenomenon, we found that the P2X2 c-terminus (CT) could interfere with P2X2 and P2X4 interactions with NMDARs. We also report that 11 distal CT residues on the P2X4 facilitate the P2X4-NMDAR interaction, and that a peptide consisting of these P2X4 CT residues (11C) can disrupt the interaction between NMDARs and P2X2 or P2X4. Collectively, these results provide new evidence for the modulatory nature of P2X2 and P2X4, suggesting they might play a more nuanced role in the CNS.

IIAEK (Ile-Ile-Ala-Glu-Lys, lactostatin) is a novel pentapeptide from bovine milk β-lactoglobulin which lowers cholesterol levels. However, the molecular mechanisms underlying the suppression of intestinal cholesterol absorption by IIAEK are unknown. Therefore, we evaluated the effects of IIAEK on intestinal cholesterol metabolism in Caco-2 cells in a human intestinal model. We found that IIAEK significantly reduced the expression of intestinal cholesterol metabolism-associated genes, particularly that of the ATP-binding cassette transporter A1 (ABCA1) protein. Subsequently, we chemically synthesized a novel molecular probe, IIXEK, which can visualize a complex of target proteins interacting with photoaffinity-labeled IIAEK by fluorescent substances. Photoaffinity labeling and MS analysis with IIXEK for the rat small intestinal mucosa and intestinal lipid raft fractions of Caco-2 cells, we identified intestinal alkaline phosphatase (IAP) as a specific molecule interacting with IIAEK and discovered IIAEK common binding amino acid sequence, GFYLFVEGGR. Transfection of IAP siRNA counteracted the decrease in ABCA1 mRNA levels in Caco-2 cells. IIAEK significantly increased IAP mRNA and protein levels, and significantly decreased ABCA1 mRNA and protein levels in Caco-2 cells. In conclusion, we found that IIAEK targets IAP to improve cholesterol metabolism via a novel signaling pathway with a specific activation of IAP and down-regulation of intestinal ABCA1.

RNA polymerase II (RNAPII) frequently transcribes non-protein coding DNA sequences in eukaryotic genomes into long non-coding RNA (lncRNA). Here, we focus on the impact of the act of lncRNA transcription on nearby functional DNA units. Distinct molecular mechanisms linked to the position of lncRNA relative to the coding gene illustrate how non-coding transcription controls gene expression. We review the biological significance of the act of lncRNA transcription on DNA processing, highlighting common themes, such as mediating cellular responses to environmental changes. This review presents the background in chromatin signaling to appreciate examples in different organisms where we can interpret functions of non-coding DNA through the act of RNAPII transcription.

Nine components (C1-C9) were isolated from chloroform fraction of fractionated methanol extracts of Vernonia amygdalina leaves (FMEVA) by column chromatography. All the components C1 to C9 were purified and screened for hypoglycaemic activities in type-2 diabetic rats. The most potent hypoglycaemic component was elucidated on the basis of extensive spectroscopic (1D-, 2D-NMR, GC-MS, FTIR) data analysis. The Component C5 was found to be the most potent hypoglycaemic in reducing blood glucose by 12.55 ± 3.55% at 4 h post-oral administration, when compared to the positive (18.07 ± 1.20%) and negative (-1.99 ± 0.43%) controls. The spectroscopic data analysis reveals that the isolated compound has a structure consistent with 11β,13-dihydrovernolide. The isolated compound is part of the hypoglycaemic components present in V. amygdalina leaves that is responsible for the anti-diabetic activities. Further research is needed in the development of this compound or its derivatives for pharmaceutical use.

Seafood is considered one of the main food allergen sources by the European Food Safety Authority (EFSA). It comprises several distinct groups of edible aquatic animals including fish and shellfish such as crustacean and mollusks. Recently the EFSA recognized the high risk of food allergy over the world and established the necessity of developing new methodologies for its control. Consequently, accurate, sensitive and fast detection methods for seafood allergy control and detection in food products are highly recommendable. In this work, we present a comprehensive review of the applications of the proteomics methodologies for the detection and quantification of seafood allergens. For that, two consecutive proteomics strategies (Discovery and Targeted Proteomics) applied for the study and control of seafood allergy are reviewed in detail. In addition, future directions and new perspectives were also provided.

In numerous instances, the fate of a single cell not only represents its peculiar outcome but also contributes to the overall status of an organism. In turn, cell division cycle and its control strongly influence cell destiny playing a critical role in targeting it towards a specific phenotype. Several factors participate to the control of growth and among them p27 and p57, two proteins modulating various transitions of cell cycle, appear to play key functions. In this review, the major features of p27 and p57 will be described, focusing in particular on their recently identified roles not directly correlated to cell cycle modulation. Then, their possible role as molecular effectors of polyphenols activities are discussed. Polyphenols represent a large family of natural bioactive molecules that have been demonstrated to play promising protective activities against several human diseases. Their use has also been proposed in association with classical therapies for ameliorating their clinical effects and for diminishing their negative side activities. The importance of p27 and p57 in polyphenol cellular effects will be discussed with the aim of identifying novel therapeutic strategies for the treatment of important human diseases, such as cancers, characterized by an altered control of growth.

Growing cells is the typical mode of operation in many aspects of biotechnology and metabolic engineering. This comes about due to cell growth processes creating a driving force that pull metabolic flux along different metabolic pathways, that indirectly help move substrate to product. But, there is an alternative mode of operation that uses resting (non-growing) cells to achieve similar or even higher productivities. In general, resting cells are provided with carbon substrates for biocatalytic reactions but starved of nitrogen or phosphorus. Such resting cells have been usefully employed in many forms of biocatalysis and biotransformation, with or without cofactor regeneration. However, much remains unknown about the transcriptome and metabolome of resting cells in biotransformation settings. This short writeup provides the backdrop of resting cells in biocatalysis, documents their use in biotransformation with application examples, and identifies research gaps that could be filled with contemporary RNA-seq and mass spectrometry proteomics technology. Overall, utility of resting cells in biocatalysis and the extant knowledge gap in their fundamental physiology are highlighted in this resource.

Recent COVID-19 outbreak has come into prominence the pathogenetic mechanisms underlying the Biology and Biochemistry of viral infections. COVID-19 illness is brought about by infection with the severe acute respiratory syndrome coronavirus SARS-CoV-2 [1,2], an enveloped positive single stranded RNA virus (ssRNA+). From a lipidomics viewpoint, there is a variety of mechanisms involving virus infection that encompass virus entry, disturbance of host cell lipid metabolism, and the role played by diverse lipids in regard to the infection effectiveness. All these aspects have currently been tackled separately as independent issues and focusing on the function of proteins. Here we review the role of cholesterol and other lipids in in ssRNA+ and SARS-COV-2 infection.

Oleuropein, glycosylated secoiridoid present in olive leaves is known to be an important antioxidant phenolic compound. We studied the antioxidant effect of low doses of oleuropein aglycone (3,4-DHPEA-EA) and oleuropein aglycone peracetylated (3,4-DHPEA-EA(P)) in murine C2C12 myocytes treated with hydrogen peroxide (H2O2). Both compounds were used at a concentration of 10 μM and were able to inhibit cell death induced by the H2O2 treatment, with 3,4-DHPEA-EA(P) being more. Under our experimental conditions, H2O2 efficiently induced the phosphorylated-active form of JNK and of its downstream target c-Jun. We demonstrated, by Western blot analysis, that 3,4-DHPEA-EA(P) was efficient in inhibiting the phospho-active form of JNK. This data suggests that the growth arrest and cell death of C2C12 proceeds via the JNK/c-Jun pathway. Moreover, we demonstrated that 3,4-DHPEA-EA(P) affects the myogenesis of C2C12 cells; because MyoD mRNA levels and the differentiation process are restored with 3,4-DHPEA-EA(P) after treatment. Overall, the results indicate that 3,4-DHPEA-EA(P) prevents ROS-mediated degenerative process in a genomic and epigenomic manner by functioning as an efficient antioxidant.

Retinoic acid receptor-related orphan receptor γ (RORγ) is a transcription factor regulating the expression of the pro-inflammatory cytokine IL-17 in human T helper 17 (Th17) cells. Activating RORγ can induce multiple IL-17-mediated autoimmune diseases but may also be useful for anticancer therapy. Its deep immunological functions make RORɣ an attractive drug target. Over 100 crystal structures have been published describing atomic interactions between RORɣ and agonists and inverse agonists. In this review, we focus on the role of dynamic properties and plasticity of the RORɣ orthosteric and allosteric binding sites by examining structural information from crystal structures and simulated models. We discuss the possible influences of allosteric ligands on the orthosteric binding site. We find that high structural plasticity favors the druggability of RORɣ, especially for allosteric ligands.

Cortical theta burst stimulation (TBS) structured as intermittent (iTBS) and continuous (cTBS) could prevent the progression of the experimental autoimmune encephalomyelitis (EAE). The interplay of brain antioxidant defense systems against overproduction of reactive oxygen, nitrogen, and thiol species induced by EAE has not been entirely investigated, just as the effect of iTBS or cTBS on oxidative-nitrogen stress (ONS) in EAE rats. Dark Agouti strain female rats were tested for the effects of EAE and TBS. The rats were randomly divided into the following groups: C - control, EAE - rats immunized for EAE, CFA - rats immunized with Complete Freund's adjuvant; iTBS and cTBS groups, and EAE+iTBS and EAE+cTBS - health and EAE rats exposed to iTBS and cTBS, respectively; EAE+iTBSsh and EAE+cTBSsh - sham stimulated EAE rats with the same noise artifacts of iTBS and cTBS, respectively. Superoxide dismutase activity, levels of superoxide anion (O2•-), lipid peroxidation, glutathione (GSH), nicotinamide adenine dinucleotide phosphate (NADPH) and thioredoxin reductase (TrxR) activity were analyzed in rat spinal cords homogenates. The severity of EAE clinical coincided with the climax of ONS, based on the increase of superoxide anion and lipid peroxidation; depletion of total thiols, GSH and NADPH; and decrease of SOD activity. The TrxR imposed the most sensitive response against the applied central nervous system (CNS) stressors to rats. We concluded that the TrxR upregulation meritoriously compensates decreased ROS sequestrating and GSH systems in EAE. Both iTBS and cTBS modulate the biochemical environment at a distance from the area of stimulation against ONS, accomplish a similar effect on TrxR activity to EAE and healthy rats, and alleviate symptoms of EAE.

Soft drinks exist in various forms and brands and are marketed by different brewery industries across the country. Nigeria been an underdeveloped country were aforesaid health problems are common, people tends to consume them lots believing it gives more energy. This study was therefore set to assess the sugar content from different soft drinks sold in Nigeria. Ten (10) soft drinks registered and licensed in Nigeria by their respective companies were purchased from a supermarket in Ihumudumu environment of Ekpoma, Nigeria. The sugar contents were assessed using standard methods. Result showed significant differences (p<0.05) in ranges of 0.36g/100ml (Limca) to 3.88g/100ml (Teem Bitter Lemon). Conclusively, the sugar contents were all below recommended value and it can be consumed by those who need low sugar in the body.

As the world endures the coronavirus disease 2019 (COVID-19) pandemic, conditions of 35 million vulnerable individuals struggling with substance use disorders (SUDs) worldwide have not received sufficient attention for their special health and medical needs. Many of these individuals are complicated by underlying health conditions, such as cardiovascular and lung diseases and undermined immune systems. During the pandemic, access to the healthcare systems and support groups is greatly diminished. Current research on COVID-19 has not addressed the unique challenges facing individuals with SUDs, including the heightened vulnerability and susceptibility to the disease. In this systematic review, we will discuss the pathogenesis and pathology of COVID-19, and highlight potential risk factors and complications to these individuals. We will also provide insights and considerations for COVID-19 treatment and prevention in patients with SUDs.

Purpose of this article is to project from the point of view of the scientific data concerning PAF and as a new approach by involving PAF, in order to contribute to clarification and proposal a possible mechanism of action of the coronavirus SARS-CoV-2 and to give a possible explanation for its observed side effects-complications of Covid-19 disease. The article is not intended to suggest any specific drugs, but clarifying the mechanism is the first step in illuminating the direction of prescribing the targets of medication that may be appropriate for the prevention, treatment, and cure of Covid-19 disease. More specifically, the purpose of this article is to provide unequivocal evidence indicating that: 1 It would not be unexpected for PAFR to participate in the entry of coronavirus SARS-CoV-2 into the cell. 2 There is interdependence and relationship of the receptors ACER and PAFR, which means the involvement of PAF in the processes related to coronavirus SARS-CoV-2 as well as to its effects exercised by the coronavirus SARS-CoV-2. 3 The receptors ACER, PAFR and TLRs are also involved in inflammation as well as in virus binding to stabilize and enter into the cell, and so they are "communicating vessels" with PAF being " a missing link" 4 Consequently there is a correlations of COVID-19 disease manifestations and biological characteristics with those caused by PAF, which shows the involvement of PAF in COVID-19 disease.

Retinoic acid receptor-related orphan receptor γ (RORγ) is a transcription factor regulating the expression of the pro-inflammatory cytokine IL-17 in human T helper 17 (Th17) cells. Activating RORγ can induce multiple IL-17-mediated autoimmune diseases but may also be useful for anticancer therapy. Its deep immunological functions make RORɣ an attractive drug target. Over 70 crystal structures have been published describing atomic interactions between RORɣ and agonists and inverse agonists. In this review, we focus on the role of dynamic properties and plasticity of the RORɣ orthosteric and allosteric binding sites by examining structural information from crystal structures and simulated models. We discuss the possible influences of allosteric ligands on the orthosteric binding site. We find that high structural plasticity favors the druggability of RORɣ, especially for allosteric ligands.

The current pneumonia epidemic could evolve into a pandemic on a global scale if not effectively contained. The COVID-19 virus possesses a 61-amino acid open reading frame resembling SARS-CoV virulence factor - ORF6 peptide. The isoleucine content is 15.9% in ORF6 of SARS-CoV versus 16.4% of that in SARS-CoV-2. Given the proton affinity in the carbonyl oxygen in isoleucine, augmented proton traffic can enhance proton-ion antiport and prompt cell swelling. Calorie restriction has been confirmed in animal studies to extend lifespan, and its underlying mechanism is not fully known. As the content of essential amino acids in the open reading frame of SARS-CoV-2 reaches 57.4%, a starch/vitamin diet served for short period of time does not give rise to essential amino acids and halts virion production, which could be adopted as prophylactic approach of many viral infections. Plant-based diet or fasting/boiled rice water can also minimize the intake of essential amino acids or all amino acids respectively. Furthermore, several proteins of SARS-CoV-2 possess high valine plus glycine content which is implicated in heart disease, justifying the aforementioned approaches.

Tree members of the genus Acacia have benefits that are obvious for enhancing soil fertility in farming, forestry and agroforestry in regions with nutrient-poor soils, and for restoring degraded ecosystems and lands. Nevertheless, the species of the genus Acacia have got the potential to bring about significant adverse impacts on biodiversity or ecosystem functioning when it gets invasive. The ecology of the species in nearly all areas of its created range remains poorly understood. Here we have compiled the information regarding the importance, cultivation and the production of important species of genus Acacia. We hope this information will be useful to get awareness about the crucial trees in the genus Acacia.

The rapidly increasing adoption of high-resolution accurate-mass methods in analytical laboratories has fueled demand for instruments that combine high performance and reliability with small size and greater ease-of-use. This paper presents the major design principles that are driving the evolution of the hybrid quadrupole-Orbitrap instrument architecture to enable a greater range of applications and users. These principles may be summarized as follows: better usage of physical space and better access for service by means of size reduction of pumping and ion optics; expanded use of technologies from electronics in ion-optical design; flexibility in performance via modularity of design of the hardware and software components; and, harmonization of interfaces with other instruments to facilitate sharing and transferability of analytical workflows. The design of a novel family of hybrid mass spectrometers is described in detail, and performance evaluation is carried out on a wide variety of samples for its three representatives: the Orbitrap Exploris 120, Orbitrap Exploris 240 and Orbitrap Exploris 480 mass spectrometers.The new instrument family is shown to offer compelling potential not only for high-end proteomics and biopharmaceutical applications, but also for screening, trace, targeted and clinical analysis by liquid chromatography/mass spectrometry methods.

Corona virus disease 2019 (COVID-19) is caused by a Severe Acute Respiratory Syndrome-Coronavirus 2 (SARS-CoV-2), which is a positive strand RNA virus. The SARS-CoV-2 genome and its association to SAR-CoV-1 vary from ca. 66% to 96% depending on the type of betacoronavirdeae family members. With several drugs, viz. chloroquine, hydroxychloroquine, ivermectin, artemisinin, remdesivir, azithromycin considered for clinical trials, there has been an inherent need to find distinctive antiviral mechanisms of these drugs. Curcumin, a natural bioactive molecule has been shown to have a therapeutic potential for various diseases, but its effect on COVID-19 has not been explored. In this study, we show the binding potential of curcumin targeted to a variety of SARS-CoV-2 proteins, viz. spike glycoproteins (PDB ID: 6VYB), nucleocapsid phosphoprotein (PDB ID: 6VYO), membrane glycoprotein (PDB ID: 6M17) along with nsp10 (PDB ID: 6W4H) and RNA dependent RNA polymerase (PDB ID: 6M71) structures. Our results indicate that curcumin has high binding affinity towards nucleocapsid and nsp 10 proteins with potential antiviral activity.

Corona virus disease 2019 (COVID-19) is caused by a Severe Acute Respiratory Syndrome-Coronavirus 2 (SARS-CoV-2), which is a positive strand RNA virus. The SARS-CoV-2 genome and its association to SAR-CoV-1 vary from ca. 66% to 96% depending on the type of betacoronavirdeae family members. With several drugs, viz. chloroquine, hydroxychloroquine, ivermectin, artemisinin, remdesivir, azithromycin considered for clinical trials, there has been an inherent need to find distinctive antiviral mechanisms of these drugs. Curcumin, a natural bioactive molecule has been shown to have a therapeutic potential for various diseases, but its effect on COVID-19 has not been explored. In this study, we show the binding potential of curcumin targeted to a variety of SARS-CoV-2 proteins, viz. spike glycoproteins (PDB ID: 6VYB), nucleocapsid phosphoprotein (PDB ID: 6VYO), membrane glycoprotein (PDB ID: 6M17) along with nsp10 (PDB ID: 6W4H) and RNA dependent RNA polymerase (PDB ID: 6M71) structures. Our results indicate that curcumin has high binding affinity towards nucleocapsid and nsp 10 proteins with potential antiviral activity.

Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) is a respiratory syndrome caused by positive RNA virus resulting in outbreak of corona virus disease 2019 (COVID-19). The SARS-CoV-2 genome and its association to SAR-CoV-1 vary from ca. 66% to 96% depending on the type of betacoronavirdeae family members. With several drugs, viz. chloroquine, hydroxychloroquine, ivermectin, quinidine, artemisinin, remdesivir, azithromycin considered for clinical trials, there has been an inherent need to find distinctive antiviral mechanisms of these drugs. On the other hand, curcumin, a natural bioactive molecule has been shown to have a therapeutic potential for various diseases, but no role of it in COVID-19 has been explored. In this work, we show the binding potential of curcumin targeted to a host of SARS-CoV-2 proteins, viz. spike glycoproteins (PDB ID: 6VYB), nucleocapsid phosphoprotein (PDB ID: 6VYO), membrane glycoprotein (PDB ID: 6M17) along with nsp10 (PDB ID: 6W4H) and RNA dependent RNA polymerase (PDB ID: 6M71) structures. Our results indicate that curcumin has potential antiviral protein binding affinity towards SARS-CoV-2 proteins which is comparable with other repurposed drugs that are considered for clinical trials.

COVID-19 is a rapidly evolving medical emergency that has drawn global attention, unprecedented in any disease of its kind in recent times. The magnitude of the health crisis emerging from this pandemic has overwhelmed health care workers worldwide and called in for extraordinary measures to contain this virus. A simple Pubmed query on “COVID-19” returned with 12214 articles (as on May 17^th, 2020), published just within a few months. A detailed survey revealed around 250 clinical reports, 8 clinical trials, 9 meta-analyses, and 906 reviews that were published during this time span. Combining the strings “COVID-19 and Pregnancy” yielded a total of 132 reports while querying “COVID-19 and Placenta” returned with just 11 articles Even taking into considerations that few materials are in the PrePrint Server, we still have a gross under-representation of studies addressing the effect of this disease on pregnancy outcome and maternal & child health. An essential aspect of a successful pregnancy is proper placentation, where transiently invasive placental trophoblast cells invade the maternal endometrium to establish a functional feto-maternal communication. Based on the elegant study by David. E. Gordon, et al. published in Nature (April 30, 2020), which identified 332 human host proteins interacting with SARS-nCoV2 using an affinity-based purification, we interrogated several gene expression data sets available at NCBI-GEO related to trophoblast invasion and differentiation. Both of these processes are indispensable for placentation and fetal survival. Our analysis showed several overlaps with the interactome proteins implying that SARS-CoV-2 infection can affect several proteins, which are crucial for trophoblasts function. GeneMANIA and STRING based functional analysis further revealed that several of that SARS-CoV-2 interacting trophoblast proteins as a hub for the protein-protein interaction network. Our study thus elucidates the possible effect of SARS-CoV-2 infection on placenta formation and pregnancy outcome.

The coevolution of species drives diversity in animals and plants and contributes to natural selection, while in host–parasite coevolution, a parasite may complete an incomplete evolutionary/developmental function by utilizing the host cell’s machinery. Analysis of related older data suggests that Plasmodium falciparum (P. falciparum), the pathogen of malaria tropica, cannot survive outside its human host because it is unable to perform the evolutionarily first protein glycosylation or blood group-independent (serologically A-like) O-GalNAcα1-Ser/Thr-R, Tn antigen (“T nouvelle”) formation owing to its inability for synthesizing the amino sugar N-acetyl-d-galactosamine (GalNAc). This parasite breaks the species barrier via hijacking the host's A-like/Tn formation by abundantly expressing serine residues and creating hybrid A-like/Tn structures. In the human blood group O(H), these hybrid structures are attacked by the germline-encoded nonimmune polyreactive immunoglobulin M (IgM), which physiologically regulates the expression of the syngeneic A-like/Tn antigen. In non-O blood groups, this antibody molecule has undergone the phenotypic accommodation of plasma proteins, which results in loss of blood group A- and B-corresponding anti-A and anti-B isoagglutinin activities. This loss allows the generation of human A- and B allele-connected hybrid epitopes and the development of life-threatening disease almost exclusively in non-O blood groups. Although malaria infection occurs regardless of the blood group, the synthesis of the blood group AB enables the strongest contact with the pathogen, and molecularly precluding any isoagglutinin activity makes this group the least protected and the smallest among the ABO blood groups. In contrast, blood group O(H) individuals have the least contact with the pathogen; they maintain the isoagglutinins, rarely develop severe disease, and survive this coevolution in an immunological balance with the pathogen as the largest blood group worldwide.

Despite great strides being achieved in improving cancer patients’ outcomes through better therapies and combinatorial treatment, several hurdles still remain due to therapy resistance, cancer recurrence and metastasis. Drug resistance, culminating in relapse and metastatic disease continue to be associated with fatal disease. Cancer stem cells (CSCs) are a subpopulation of cancer cells known to be resistant to therapy and cause metastasis. Whilst the debate on whether CSCs are the origins of the primary tumor rages on, CSCs have been further characterised in many cancers with data illustrating that CSCs display great abilities to self-renew, withstand therapies due to enhanced epithelial to mesenchymal (EMT) properties, enhanced expression of ABC membrane transporters, activation of several survival signaling pathways and increased immune evasion DNA repair mechanisms. CSCs also display great heterogeneity with the consequential lack of specific CSC markers presenting a great challenge to their targeting. In this updated review we re-visit CSCs within the tumor microenvironment (TME) and present novel treatment strategies targeting CSCs. These promising strategies include targeting CSCs-specific properties using small molecule inhibitors, immunotherapy, microRNA mediated inhibitors, epigenetic methods as well as targeting CSC niche-microenvironmental factors and differentiation. Lastly, we present recent clinical trials undertaken to try to turn the tide against cancer by targeting CSC-associated drug resistance and metastasis.

Although metastasis is the primary cause of death on patients with malignant solid tumors, efficient antimetastatic therapies are not clinically available thus far. Sulfated glycosaminoglycans from marine sources have shown promising pharmacological effects, acting in different steps of the metastatic process. Oversulfated dermatan sulfate from ascidians is effective in preventing metastasis by inhibition of P-selectin, a platelet surface protein involved in the platelet-tumor cell emboli formation. We report in this work that the heparan sulfate isolated from the viscera of the ascidian Phallusia nigra drastically attenuates metastasis of colon carcinoma cells in mice. Our in vitro and in vivo assessments demonstrate that the P. nigra glycan has very low anticoagulant and antithrombotic activities and a reduced hypotension potential, although efficiently preventing metastasis. Therefore, it may be a promising candidate for the development of a novel anti-metastatic drug.

The MeshCODE framework outlined here represents a unifying theory of data storage in animals, providing read/write storage of both dynamic and persistent information in a binary format. Mechanosensitive proteins, that contain force-dependent switches, can store information persistently which can be written/updated using small changes in mechanical force. These mechanosensitive proteins, such as talin, scaffold each and every synapse creating a meshwork of switches that forms a code, a MeshCODE. Synaptic transmission and action potential spike trains would operate the cytoskeletal machinery to write and update the synaptic MeshCODEs, propagating this coding throughout the brain and to the entire organism. Based on established biophysical principles, a mechanical basis for memory provides a physical location for data storage in the brain. Furthermore, the conversion and storage of sensory and temporal inputs into a binary format identifies an addressable read/write memory system supporting the view of the mind as an organic supercomputer.

Solid tumors display complex biology and most therapies including chemotherapy cannot prevent therapy resistance and relapse. Most therapeutics target cancer cells, but recent data suggest the presence of cancer stem cells as cells with self-renewal and tumorigenic abilities. Cancer stem cell markers have been suggested to have prognostic value and can be targeted during cancer treatment and in resistant disease. CSCs have been postulated to play significant contextual roles in tumor initiation, progression, therapy resistance and metastasis. CSCs have thus been targeted by new generation cancer drugs. The transcriptional expression of several CSC markers in different cancers was evaluated by searching publicly available The Cancer Genome Atlas (TCGA) and Gene Expression Profiling Interactive Analysis (GEPIA) databases. We report here new findings on expression and prognostic significance of CSC markers in several cancers by examining the expression of CSCs markers in tumor tissues versus the adjacent normal tissues. We found that CSC markers were mostly highly expressed various tumors such as colon, lung, pancreatic and esophageal cancers. No CSC marker is expressed in the same pattern in all cancers and individual CSC marker expression was not linked to patient survival. This analysis calls for continued research on CSCs and clinical evaluation of the CSC markers in relation to prognosis of cancers in large population samples. Novel cancer drugs ought to target CSCs, cancer cells and tumor microenvironment variations.

This paper aims determine the efficacy of Kamias (Averrhoa bilimbi) fruit as a ripening agent for Cavendish banana (Musa acuminata). A quantitative experimental research design was employed in the study. Unripe Cavendish bananas and Kamias fruits were procured from the local market and the fruits were extracted to three different concentrations. Calcium carbide was used as positive control. Six bunches of unripe bananas were allowed to ripe and labeled according to the type of treatment. Ripe bananas were then subjected to sensory evaluation, titratable acidity and Benedict’s tests. Results showed that the use of Kamias fruit allowed ripening of banana for 76 hours while a 25-75% concentration of Kamias fruit extract allowed ripening for 76-96 hours. The bananas treated with Kamias Extract 75% had the highest level of acceptability and titratable acidity while the bananas treated with Kamias fruit had the highest level of reducing sugar. One-Way MANOVA reported that there is a significant difference in the duration of ripening, level of acceptability, titratable acidity and level of reducing sugar when treated with various ripening agents (p<0.05).

Almond (Prunus dulcis (Mill.) D.A.Webb) is one of the largest nut crops in the world. Recently, phenolic compounds, mostly stored in almond skin, have been associated with much of the health-promoting behavior associated with their intake. The almond skin enriched fraction obtained from cold-pressed oil residues of the endemic Moroccan Beldi ecotypes is particularly rich in chlorogenic acid. In this study, both almond skin extract (AE) and chlorogenic acid (CHL) supplements, similar to traditional positive control resveratrol, significantly increased the replicative life-span of yeast compared to the untreated group. Our results showed that AE and CHL significantly reduced the production of reactive oxygen and nitrogen species (ROS/RNS), most likely due to their ability to maintain mitochondrial function during aging, as indicated by the maintenance of normal mitochondrial membrane potential in treated groups. This may be associated with the observed activation of the anti-oxidative stress response in treated yeast, which results in activation at both gene expression and enzymatic activity levels for SOD2 and SIR2, the latter being an upstream inducer of SOD2 expression. Interestingly, the differential gene expression induction of mitochondrial SOD2 gene at the expense of the cytosolic SOD1 gene confirms the key role of mitochondrial function in this regulation. Furthermore, AE and CHL have contributed to the survival of yeast under UV-C-induced oxidative stress, by reducing the development of ROS / RNS, resulting in a significant reduction in cellular oxidative damage as evidenced by decreased membrane lipid peroxidation, protein carbonyl content and 8-oxo-guanine formation in DNA. Together, these results demonstrate the interest of AE and CHL as new regulators in the replicative life-span and control of the oxidative stress response of yeast.

A big race for the search for novel lead has begun due to the emergence of COVID-19 across the globe. More than 6,00,000 cases of afflicted patients worldwide has been reported till date with high mortality and morbidity. At present no approved drugs are known for COVID-19. Phylogenetic analysis present strong nucleotide sequence similarity of around 80% with SARS-CoV. Therefore, the drugs used for treating SARS-CoV and MERS are being used for SARS-CoV-2 also. Recently, the crystal structure of COVID-19 is reported and hence, we have used this tom predict the binding affinity with SARS-CoV-2-main protease and prepared a pharmacophore that may be used for future design of novel inhibitors.

Retinal pigment epithelial (RPE) cells are the major cell type in the epi- or sub-retinal membranes in the pathogenesis of proliferative vitreoretinopathy (PVR), which is a blinding fibrotic eye disease and still short of effective medicine. The purpose of this study is to demonstrate if Chalocomoracin (CMR), a novel purified compound from fungus-infected mulberry leaves, is able to inhibit vitreous-induced signaling events and cellular responses intrinsic to PVR. Our studies have revealed that the CMR IC50 for ARPE-19 cells is 35.5 μM at 72 hours, and that 5 μM CMR inhibits vitreous-induced Akt activation and p53 suppression; in addition we have discovered that this chemical effectively blocks vitreous-stimulated proliferation, migration and contraction of ARPE-19 cells, suggesting that CMR is a promising PVR prophylactic.

Recent clinical SARS-CoV-2 studies link diabetes, cardiovascular disease, and hypertension to increased disease severity. In the US, racial and ethnic minorities and low socioeconomic status (SES) individuals are more likely to have increased rates of these comorbidities, lower baseline health, limited access to care, increased perceived discrimination, and limited resources, all of which increase their vulnerability to severe disease and poor health outcomes from SARS-CoV-2. Previous studies demonstrated the disproportionate impact of pandemic and seasonal influenza on these populations, due to these risk factors. This paper reviews increased health risks and documented health disparities of racial and ethnic minorities and low SES individuals in the US. Pandemic response must prioritize these marginalized communities to minimize the negative, disproportionate impacts of SARS-CoV-2 on them and manage spread throughout the entire population. This paper concludes with recommendations applicable to healthcare facilities and public officials at various government levels.

The delivery of Cytochrome c (Cyt c) to the cytosol stimulates apoptosis in cells were its release from mitochondria and apoptosis induction is inhibited. We developed a drug delivery system consisting of Cyt c nanoparticles decorated with folate-poly(ethylene glycol)-poly(lactic-co-glycolic acid)-thiol (FA-PEG-PLGA-SH) to deliver Cyt c into cancer cells and test their targeting in the Lewis Lung Carcinoma (LLC) mouse model. Cyt c-PLGA-PEG-FA nanoparticles (NPs) of 253 ± 55 and 354 ± 11 nm were obtained by Cyt c nanoprecipitation, followed by surface decoration with the co-polymer SH-PLGA-PEG-FA, and compared to a nanoparticle-free formulation. Overexpression of FA in LLC cells and internalization of Cyt c-PLGA-PEG-FA nanoparticles (NPs) was confirmed by confocal microscopy. Caspase activation assays show NPs retain 88-96% Cyt c activity. The NP formulations were more efficient in decreasing LLC cell viability than the NP-free formulation, with IC50: 49.2 to 70.1 μg/ml versus 129.5 μg/ml, respectively. Our NP system is thrice as selective towards cancerous than normal cells. In-vivo studies using tagged nanoparticles show accumulation in mouse LLC tumor 5 min post-injection. In conclusion, our NP delivery system for Cyt c shows superiority over the NP-free formulation and reaches a folic acid-overexpressing tumor in an immune-competent animal model.

Covid-19 infections are rapidly spreading worldwide with more than 100000 death and thus understanding the molecular mechanism of tropism of human cells is an urgent need for drug design. We have described here a bioinformatics approach to predict the functional aspects of non-structural nsp16 protein of Corona virus. The covid-19 7098 AA large polyprotein was degraded into sixteen proteins and last nsp16 protein was found an RlmE type rRNA methyltransferase. Nsp16 has no similarity to bacterial RlmABCD but has 25 percent similarity to the bacterial RlmE protein which methylates the U2551 2-hydroxy group of Ribose. The nsp16 proteins of different corona viruses like covid-19, bat-coronavirus, SARS and MERS have strong homology. Mrm2 and Dim1 like yeast and mammalian rRNA methyltransferases have 26-33 percent homologies but not with 2-O-capping MTase as reported previously. Rrp8 MTases also has no similarity to nsp16. We postulated that mitochondrial rRNA methylation of bronchial cells were mediated by the nsp16 protein causing inhibition of protein synthesis due to poor assembly of aminoacyl-tRNA or mRNA and peptidyl transferase at the PTC. This is one of the new molecular mechanism of corona virus cellular tropism and different than ACE-2 mediated blockage of cellular signalling to inhibit aldesterone biosynthesis with abnormal Na⁺ ions in cells. We also designed primers based on nsp16 cDNA sequence (nt 20659-21552, accession no MT121215) specific for Covid-19 diagnosis by RT-PCR.

Even 20 years after the first crystal structures of [FeFe]-hydrogenase have been published, several aspects of biological hydrogen turnover are heatedly discussed. In this perspective, we give an overview on how the diversity of naturally occurring and artificially prepared, semi-synthetic [FeFe]-hydrogenases deepens our understanding of hydrogenase chemistry. In parallel, we cover new results from biophysical techniques that go beyond the scope of conventional electrochemistry, X-ray diffraction, EPR, and FTIR spectroscopy. Taking into account both proton transfer and electron transfer as well as the notorious sensitivity of [FeFe]-hydrogenases towards carbon monoxide, the discussion further touches upon the molecular proceedings of biological hydrogen turnover.

Corona Virus Infectious Disease-2019 (COVID-19) outbreak originated recently at Wuhan, China in December 2019. It has already spread rapidly to more than 200 countries and has been declared a pandemic by WHO. It is caused by a beta-coronavirus named as SARS-CoV-2. There is no definitive cure, either drug or vaccine, to treat or prevent this viral disease. Recently, the crystal structure of the main protease M^pro has been determined. M^pro is responsible for the proteolytic maturation of the polyprotein essential for the viral replication and transcription, which makes it an important drug target. The discovery of new drug molecules may take years before getting to the clinics. So, considering urgency we performed molecular docking studies using FDA approved drugs to identify molecules that could potentially bind to the substrate-binding site and inhibit SARS-CoV-2 main protease (M^pro). We used the Glide module in Schrodinger software suite to perform molecular docking studies followed by MM-GBSA based energy calculations to score the hit molecules. Molecular docking and manual analysis suggest that several drugs may bind and potentially inhibit M^pro. We also performed molecular simulations studies for selected compounds to evaluate protein-drug interactions. Interestingly, we observed only one antiviral compound, Adefovir, in the top50 list of compounds. Considering bioavailability, lesser toxicity, route of administration some of the top-ranked drugs including lumefantrine (antimalarial), dipyridamole (coronary vasodilator), dihydroergotamine (used for treating migraine), hexoprenaline (anti- asthmatic), riboflavin (vitamin B2) and pantethine (vitamin B5) may be taken forward for further in vitro and in vivo experiments to investigate their therapeutic potential.

In E. coli DNA replication forks stall on average once per cell cycle. When this occurs, replisome components disengage from the DNA, exposing an intact, or nearly intact fork. Consequently, the fork structure must be regressed away from the initial impediment so repair can occur. Regression is catalyzed by the powerful, monomeric DNA helicase, RecG. During this reaction, the enzyme couples unwinding of fork arms to rewinding of duplex DNA resulting in the formation of a Holliday junction. RecG works against large opposing forces enabling it to clear the fork of bound proteins. Following subsequent processing of the extruded junction, the PriA helicase mediates reloading of the replicative helicase DnaB leading to the resumption of DNA replication. The single-strand binding protein (SSB) plays a key role in mediating PriA and RecG functions at forks. It binds to each enzyme via linker/OB-fold interactions and controls fork loading sites in a substrate-dependent manner that involves helicase remodeling. Finally, it is displaced by RecG during fork regression. The intimate and dynamic SSB-helicase interactions play key roles in ensuring fork regression and DNA replication restart.

Lemon pectin extracted along with water-soluble flavonoids and other phytochemicals from citrus industry’s waste lemon peel via hydrodynamic cavitation in water, directly at pre-industrial scale and further isolated via freeze drying, shows exceptionally high antioxidant and non-cytotoxic activity. Preliminary investigation indicates also significant antimicrobial activity. These findings open the route to the development of new nutraceutical and healthcare application of a versatile biopolymer endowed with new functionality, rapidly and conveniently obtained from an abundant by-product of the agrofood industry.

The recent outbreak of novel coronavirus (SARS-CoV-2 or 2019-nCoV) and its spread to the whole world is currently posing one of the major threats to human health and the world economy. It has been suggested that SARS-CoV-2 is similar to SARS-CoV based on the genome sequence comparison. Despite the genomic similarity between SARS-CoV and SARS-CoV-2, the spike glycoprotein and receptor binding domain in SARS-CoV-2 shows considerable difference compared to SARS-CoV, due to the presence of several point mutations. We analyzed the receptor binding domain (RBD) from recently published 3D structure of spike glycoprotein of SARS-CoV-2 and compared with RBD of SARS-CoV. The observations highlight few important features of RBD in the light of the recently published findings from the 3D structures of spike glycoprotein and its complex with human angiotensin-converting enzyme 2 (ACE2) (Yan, R., et al. (2020); Wrapp, D., et al. (2020); Walls, A. C., et al. (2020)).

The current pneumonia epidemic could evolve into a pandemic on a global scale if not effectively contained. The COVID-19 possesses a 61-amino acid open reading frame resembling SARS-CoV virulence factor - ORF6 peptide. The isoleucine content is 15.9% in ORF6 of SARS-CoV versus 16.4% of that in COVID-19. Given the proton affinity in the carbonyl oxygen in isoleucine, augmented proton traffic can enhance proton-ion antiport and prompt cell swelling. Calorie restriction has been confirmed in animal studies to extend lifespan, and its underlying mechanism is not fully known. As the content of essential amino acids in the open reading frame of COVID-19 reaches 57.4%, a starch/vitamin diet served for short period of time does not give rise to essential amino acids and halts virion production, which could be adopted as prophylactic approach of many viral infections. Plant-based diet or fasting/boiled rice water can also minimize the intake of essential amino acids or all amino acids respectively. Furthermore, several proteins of COVID-19 possess high valine plus glycine content which is implicated in heart disease, justifying the aforementioned approaches.

The recent outbreak of novel coronavirus (2019-nCoV or SARS-CoV-2) and its spread to the whole world is currently posing one of the major threat to human health and the world economy. It has been suggested that 2019-nCoV is similar to SARS-CoV based on the genome sequence comparison. Despite the genomic similarity between SARS-CoV and SARS-CoV-2, the spike glycoprotein and receptor binding domain in SARS-CoV-2 shows considerable difference compared to SARS-CoV, due to the presence of several point mutations. We analyzed the receptor binding domain (RBD) from recently published 3D structure of spike glycoprotein of SARS-CoV-2 and compared with RBD of SARS-CoV. The observations highlight few important features of RBD in the light of the recently published findings from the 3D structures of spike glycoprotein and its complex with human angiotensin-converting enzyme 2 (ACE2) (Yan, R., et al. (2020); Wrapp, D., et al. (2020); Walls, A. C., et al. (2020)).

The Warburg effect refers to the phenomenon that cancer cells produce energy via glycolysis instead of cellular respiration. Glycolysis generated no net protons. The Warburg effect may be malignant cells’ built-in mechanism to antagonize the buildup of protons via Krebs cycle and other pathways with compromised cellular respiration. Data described in this study indicated that cancer cells were less sensitive to the presence of oxalate than non-cancer model cell lines 16HBE14o- and HaCaT. Malignant cells may resort on organic acids such as oxalate and their calcium salts to antagonize strong acids. This experiment sheds light on the role of Warburg effect in cancer cell metabolism and homeostasis.

The current pneumonia epidemic in China could evolve into a pandemic on a global scale if not effectively contained. The SARS-CoV-2 possesses a 61-amino acid open reading frame resembling SARS-CoV virulence factor - ORF6 peptide. The isoleucine content is 15.9% in ORF6 of SARS-CoV versus 16.4% of that in SARS-CoV-2. Given the proton affinity in the carbonyl oxygen in isoleucine, augmented proton traffic can enhance proton-ion antiport and prompt cell swelling. Calorie restriction has been confirmed in animal studies to extend lifespan, and its underlying mechanism is not fully known. As the content of essential amino acids in the open reading frame of SARS-CoV-2 reaches 57.4%, a starch/vitamin diet served for short period of time does not give rise to essential amino acids and halts virion production, which could be adopted as prophylactic approach of many viral infections. Plant-based diet or fasting/boiled rice water can also minimize the intake of essential amino acids or all amino acids respectively. Furthermore, several proteins of SARS-CoV-2 possess high valine plus glycine content which is implicated in heart disease, justifying the aforementioned approaches.

Lemon pectin extracted along with water-soluble flavonoids and other phytochemicals from citrus industry’s waste lemon peel via hydrodynamic cavitation in water, directly at pre-industrial scale and further isolated via freeze drying, shows exceptionally high antioxidant and non-cytotoxic activity. Preliminary investigation indicates also significant antimicrobial activity. These findings open the route to the development of new nutraceutical and healthcare application of a versatile biopolymer endowed with new functionality, rapidly and conveniently obtained from an abundant by-product of the agrofood industry.

The intermediate filament protein vimentin constitutes a critical sensor for electrophilic and oxidative stress. We previously showed that vimentin interacts with zinc, which affects its assembly and redox sensing. Here we have used vimentin wt and C328S, an oxidation-resistant mutant showing improved NaCl-induced polymerization, to assess the impact of zinc on soluble and polymerized vimentin by light scattering and electron microscopy. Zinc acts as a switch, reversibly inducing the formation of vimentin oligomeric species. High zinc concentrations elicit optically-detectable vimentin structures with a characteristic morphology depending on the support. These effects also occur in vimentin C328S, but are not mimicked by magnesium. Treatment of vimentin with micromolar zinc induces fibril-like particles that do not assemble into filaments, but form aggregates upon subsequent addition of NaCl. In contrast, when added to NaCl-polymerized vimentin, zinc increases the diameter or induces lateral association of vimentin wt filaments. Remarkably, these effects are absent or attenuated in vimentin C328S filaments. Therefore, the zinc-vimentin interaction depends on the chemical environment and on the assembly state of the protein, leading to atypical polymerization of soluble vimentin, likely through electrostatic interactions, or to broadening and lateral association of preformed filaments through mechanisms requiring the cysteine residue. Thus, impact of zinc on vimentin assembly and redox regulation is envisaged.

The aim of the study was to analyze the in ovo injection of chemically and biologically synthesized silver nano-particles (Ag NPs) using Brassica oleracea L. var capitate f. Rubra, (BOL) conjugation with L-Arginine (L-Arg) on the immune, muscle growth, survivability and hatchability of the broiler chickens. L-Arg (100 μg) conjugated with 1000 µg of Ag NPs synthesized by (BOL)-extract and L-Arg (100 μg) conjugated with 100 µg of Ag NPs chemically synthesized were injected into fertile eggs at 8d, 14d and 18d of incubation. Living embryo and hatched chicks were calculated. Survivability and hatchability were not affected by the injected dose of L-Arg (100 μg) with 1000 µg (BOL-Ag NPs) and L-Arg (100 μg) with 100 µg (C-Ag NPs) but it significantly improved when the eggs were injected on day 14 of incubation compared with those injected on days 8 or 18. Moreover, the protein expression of muscle development markers such as myogenin and myoD were significantly up-related in 14 d of incubation whereas the heat shock proteins (HSPs) such as HSP-60 and HSP-70 were significantly up-regulated in 18 d incubation. In addition, the in ovo injection on 18 d significantly increased the serum glutamate oxaloacetate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) as well the immunoglobulin (IgM) levels were increased in 14d incubation period in serum at the same concentration.

On the primitive Earth, both L- and D-amino acids would have been present. However, only L-amino acids are essential blocks to construct proteins in modern life. To study the relative stability of homochiral and heterochiral peptides, a variety of computational methods were employed. 10 prebiotic amino acids (Gly, Ala, Asp, Glu, Ile, Leu, Pro, Ser, Thr, and Val) were previously determined by multiple previous meteorite, spark discharge, and hydrothermal vent studies. We focused on what had been reported as primary early Earth polypeptide analogs: 1ARK, 1PPT, 1ZFI, and 2LZE. Tripeptide composed of only Asp, Ser, and Val exemplified that different positions (i.e., N-terminus, C-terminus, and middle) made a difference in minimal folding energy of peptides, while the classification of amino acid (hydrophobic, acidic, or hydroxylic) did not show significant difference. Hierarchical cluster analysis for dipeptides with all possible combinations of the proposed 10 prebiotic amino acids and their D-amino acid substituted derivatives generated five clusters. Prebiotic polypeptides were built up to test the significance of molecular fluctuations, secondary structure occupancies, and folding energy differences based on these clusters. Most interestingly, among 129 residues, mutation sensitivity profiles presented that the ratio of more stable to less stable to equally stable D-amino acids was about 1:1:1. In conclusion, some combinations of a mixture of L- and D-amino acids can act as essential building blocks of life. Peptides with α-helices, long β-sheets, and long loops are usually less sensitive to D-amino acid replacements in comparison to short β-sheets.

The current pneumonia epidemic in China could evolve into a pandemic on a global scale if not effectively contained. The 2019-nCoV possesses a 61-amino acid open reading frame resembling SARS-CoV virulence factor - ORF6 peptide. The isoleucine content is 15.9% in ORF6 of SARS-CoV versus 16.4% of that in 2019-nCoV. Given the proton affinity in the carbonyl oxygen in isoleucine, augmented proton traffic can enhance proton-ion antiport and prompt cell swelling. As the content of essential amino acids in the open reading frame of 2019-nCoV reaches 57.4%, a starch/vitamin diet served for short period of time does not give rise to essential amino acids and halts virion production, which could be adopted as prophylactic approach of many viral infections. Plant-based diet or fasting/boiled rice water can also minimize the intake of essential amino acids or all amino acids respectively. Calorie restriction has been confirmed in animal studies to extend lifespan, and its underlying mechanism is not fully known. Furthermore, several proteins of 2019-nCoV possess high valine plus glycine content, which is implicated in heart disease.

Autophagy is an essential mechanism to maintain cellular homeostasis. Besides its role in controlling the quality of cytoplasmic components, it participates in nutrient obtaining and lipid mobilization under stressful conditions. Furthermore, autophagy is involved in the regulation of systemic metabolic, a function mainly performed by neuronal populations of the arcuate nucleus of the hypothalamus. Several studies have shown that blockade of autophagy in these neurons can affect central regulation of metabolism and impact body energy balance. Moreover, hypothalamic autophagy can be altered during obesity. However, neurons are not the only cell type involved in the central regulation of metabolism. Astrocytes, essential cells for brain homeostasis, are key metabolic regulators. They can sense metabolic signals in the hypothalamus and modulate systemic functions as glucose homeostasis and feeding response. Moreover, the response of astrocytes to obesity has been widely studied. Astrocytes are important mediators of brain inflammation and can be affected by increased levels of saturated fatty acids associated to obesity. Although autophagy plays important roles for astrocyte homeostasis and functioning, the contribution of astrocyte autophagy to systemic metabolism has not been analysed. Furthermore, how obesity can impact astrocyte autophagy is poorly understood. More studies are needed in other to understand the contribution of astrocyte autophagy to metabolism.

It has been long recognized that under hypoxia conditions cancer cells reprogram their metabolism through shift from oxidative phosphorylation (OXPHOS) to glycolysis to meet elevated requirements in energy and nutrients for proliferation, migration and survival. However, data accumulated over the last years increasingly evidence that cancer cells can revert from glycolysis to OXPHOS and maintain both reprogrammed and oxidative metabolism even in the same tumor. The phenomenon denoted as cancer cell metabolic plasticity or hybrid metabolism depends on a tumor micro-environment, which is highly heterogeneous and influenced by intensity of vasculature and blood flow, oxygen concentration, nutrient and energy supply, and requires regulatory interplay between multiple oncogenes, transcription factors, growth factors, reactive oxygen species (ROS), etc. Hypoxia-inducible factor-1 (HIF-1) and AMP-activated protein kinase (AMPK) represent key modulators of switch between reprogrammed and oxidative metabolism. The present review focuses on cross-talks between HIF-1, GLUTs, and AMPK and other regulatory proteins including oncogenes such as c-Myc, p53 and KRAS, growth factor-initiated PKB/Akt, PI3K and mTOR signaling pathways and tumor suppressors such as LKB1 and TSC1 in controlling cancer cell metabolism. The multiple switches between metabolic pathways can underlie chemo-resistance to conventional anti-cancer therapy and should be taken into account in choosing molecular targets to discovery novel anti-cancer drugs.

Besides their primary involvement in the recycling and degradation of proteins in endo-lysosomal compartments but also in specialized biological functions, cysteine cathepsins are pivotal proteolytic contributors of various deleterious diseases. While the molecular mechanisms of regulation by their natural inhibitors have been exhaustively studied, less is currently known about how their enzymatic activity is modulated during the redox imbalance associated with an oxidative stress and their exposure resistance to oxidants. More specifically, there is only patchy information on the regulation of lung cysteine cathepsins, while the respiratory system is directly exposed to countless exogenous oxidants contained in dust, tobacco, combustion fumes, and industrial or domestic particles. Papain-like enzymes (clan CA, family C1, subfamily C1A) encompass a conserved catalytic thiolate-imidazolium pair (Cys25-His159) in their active site. Despite the sulfhydryl group (with a low acidic pKa) is a potent nucleophile highly susceptible to chemical modifications, some cysteine cathepsins reveal an unanticipated resistance to oxidative stress. Beside an introductory chapter and a peculiar attention to lung cysteine cathepsins, the purpose of this review is to afford a concise update of the current knowledge on molecular mechanisms associated to the regulation of cysteine cathepsins by redox balance and by oxidants (e.g. Michael acceptors, reactive oxygen and nitrogen species).

The current pneumonia epidemic in China could evolve into a pandemic on a global scale if not effectively contained. The 2019-nCoV possesses a 61-amino acid open reading frame resembling SARS-CoV virulence factor - ORF6 peptide. The isoleucine content is 15.9% in ORF6 of SARS-CoV versus 16.4% of that in 2019-nCoV. Given the proton affinity in the carbonyl oxygen in isoleucine, augmented proton traffic can enhance proton-ion antiport and prompt cell swelling. As the content of essential amino acids in the open reading frame of 2019-nCoV reaches 57.4%, a starch/vitamin diet served for short period of time does not give rise to essential amino acids and halts virion production, which could be adopted as prophylactic approach of many viral infections. Plant-based diet or fasting/boiled rice water can also minimize the intake of essential amino acids or all amino acids respectively. Calorie restriction has been confirmed in animal studies to extend lifespan, and its underlying mechanism is not fully known. Furthermore, several proteins of 2019-nCoV possess high valine plus glycine content, which is implicated in heart disease.

Single molecule de novo protein sequencing based on the 'superspecificity' of amino-acyl tRNA synthetases (aaRS) is proposed. An unfolded protein molecule is threaded through a nanopore in an electrolytic cell (e-cell) to expose the terminal residue in the e-cell's trans chamber. After the residue is cleaved with an exopeptidase, a set of tRNAs, their aaRSs, and ATP are added to trans. An aaRS charges a cognate tRNA molecule with the residue. The charged tRNA (along with the other reactants) is transferred to an extended e-cell with N (20 ≤ N ≤ 61) pores in N individual cis chambers and a single trans chamber. Each pore holds an RNA molecule ending in a unique codon that is exposed in trans. The charged tRNA's anticodon base-pairs with the terminal codon of an RNA. If tRNAs and residues are fluorescently tagged with two different colors, the residue can be identified from the observed position of the resulting color pair. As charging is 'superspecific' identification is unambiguous. The protein molecule in the first e-cell is advanced by one residue and the process repeated. In this approach there is no need for precise pore current or optical intensity measurements. Potential implementation issues are discussed. Other possibilities, including one in which the terminal residue is cleaved after charging, are also examined.

In present study, the effect of Inonotus Obliquus extracts used in traditional medicine was investigated on the expression of matrix metalloproteinases-1 (MMP-1) in the normal human dermal fibroblasts. As shown our results, extracts of Inonotus Obliquus decreased MMP1 expression in oxidative stress-exposed normal human dermal fibroblasts. Additionally, Inonotus Obliquus extracts decreased AP-1 transcriptional activity and phospho-JNK in oxidative stress exposed normal human dermal fibroblasts. The results suggest that Inonotus Obliquus extracts decreased MMP1 expression using decreasing AP-1 transcriptional activity and phospho-JNK. Therefore, Inonotus Obliquus extracts has potential to reduce formation of wrinkle and to use as a cosmetic ingredient.

Manipulating autophagy is a promising strategy for treating cancer as several autophagy inhibitors shown to induce autophagic cell death. One of these, autophagonizer (APZ), induces apoptosis-independent cell death by binding an unknown target via an unknown mechanism. To identify APZ targets we used a label-free drug affinity responsive target stability (DARTS) approach with a liquid chromatography/tandem mass spectrometry (LC-MS/MS) readout. Of 35 protein interactors, we identified Hsp70 as a key target protein of unmodified APZ in autophagy. Either APZ treatment or Hsp70 inhibition attenuates integrity of lysosomes, which leads to autophagic cell death exhibiting an excellent synergism with a clinical drug, temozolomide, in vitro, in vivo, and orthotropic glioma xenograft model. These findings demonstrate the potential of APZ to induce autophagic cell death and its development to combinational chemotherapeutic agent for glioma treatment. Collectively, our study demonstrated that APZ, a new autophagy inhibitor, can be used as a potent antitumor drug candidate to get over unassailable glioma and revealed a novel function of Hsp70 in lysosomal integrity regulation of autophagy.

Acidic glycosphingolipids, gangliosides are highly and consistently expressed in nervous tissues of vertebrates at high levels. Therefore, they have been believed to be largely involved in the development and function of nervous systems. Recent studies with genetic engineering of glycosyltransferase genes have revealed novel aspects of roles of gangliosides in the regulation of nervous tissues with substantial basis. In this review, novel findings on the ganglioside functions and their modes of action elucidated mainly by studies of gene knockout mice have been summarized. In particular, roles of gangliosides in the regulation of lipid rafts to maintain the integrity of nervous systems have been reported with a focus on the roles in the regulation of neuro-inflammation and neurodegeneration via complement systems. In addition, recent advances in the studies of congenital neurodisorders due to the genetic mutaions of ganglioside synthase genes, and also in the techniques for the analysis of ganglioside functions have been introduced.

Although natural products are an important source of drugs and drug leads, identification and validation of their target proteins have proven difficult. Here, we report the development of a systematic strategy for target identification and validation employing drug affinity responsive target stability (DARTS) and mass spectrometry imaging (MSI) without modifying or labeling natural compounds. Through a validation step using curcumin, which targets aminopeptidase N (APN), we successfully standardized the systematic strategy. Using label-free voacangine, an antiangiogenic alkaloid molecule as the model natural compound, DARTS analysis revealed vascular endothelial growth factor receptor 2 (VEGFR2) as a target protein. Voacangine inhibits VEGFR2 kinase activity and its downstream signaling by binding to the kinase domain of VEGFR2, as was revealed by docking simulation. Through cell culture assays, voacangine was found to inhibit the growth of glioblastoma cells expressing high levels of VEGFR2. Specific localization of voacangine to tumor compartments in a glioblastoma xenograft mouse was revealed by MSI analysis. The overlap of histological images with the MSI signals for voacangine was intense in the tumor regions and showed colocalization of voacangine and VEGFR2 in the tumor tissues by immunofluorescence analysis of VEGFR2. The strategy employing DARTS and MSI to identify and validate the targets of a natural compound as demonstrated for voacangine in this study is expected to streamline the general approach of drug discovery and validation using other biomolecules including natural products.

Myelin ensheathes selected axonal segments within the nervous system, resulting primarily in nerve impulse acceleration, as well as mechanical and trophic support for neurons. In the central and peripheral nervous systems, various proteins that contribute to the formation and stability of myelin are present, which also harbour pathophysiological roles in myelin disease. Many myelin proteins share common attributes, including small size, high hydrophobicity, multifunctionality, longevity, and intrinsic disorder. With recent advances in protein biophysical characterization and bioinformatics, it has become evident that intrinsically disordered proteins (IDPs) are abundant in myelin, and their flexible nature enables multifunctionality. Here, we review known myelin IDPs, their conservation, molecular characteristics and functions, and their disease relevance, along with open questions and speculations. We place emphasis on classifying the molecular details of IDPs in myelin and correlate these with their various functions, including susceptibility to post-translational modifications, function in protein-protein and protein-membrane interactions, as well as their role as extended entropic chains. We discuss how myelin pathology can relate to IDPs and which molecular factors are potentially involved.

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.

The MAP kinase ERK5 contains an N-terminal kinase domain and a unique C-terminal tail including a nuclear localization signal and a transcriptional activation domain. ERK5 is activated in response to growth factors and stresses, and regulates transcription at the nucleus by either phosphorylation or interaction with transcription factors. MEK5-ERK5 pathway plays an important role regulating cancer cell proliferation and survival. Therefore, it is important to define the precise molecular mechanisms implicated in ERK5 nucleo-cytoplasmic shuttling. We previously described that the molecular chaperone Hsp90 stabilizes and anchors ERK5 at the cytosol, and that ERK5 nuclear shuttling requires Hsp90 dissociation. Here, we show that MEK5 or Cdc37 overexpression -mechanisms that induce nuclear ERK5- induced ERK5 SUMO-2 modification at residues Lys6/Lys22 in cancer cells. We also show that overexpression of the SUMO protease SENP2 completely abolished endogenous ERK5 nuclear localization in response to EGF stimulation. Furthermore, mutation of these SUMO sites abolished the ability of ERK5 to translocate to the nucleus and to promote prostatic cancer PC-3 cell proliferation. These results allow us to propose a more precise mechanism: in response to MEK5 activation, ERK5 SUMOylation favors the dissociation of Hsp90 from the complex, allowing ERK5 nuclear shuttling and activation of transcription.

A new pathway leading to the n-10 fatty acid series has been recently evidenced, starting from sapienic acid - a monounsaturated fatty acid (MUFA) resulting from the transformation of palmitic acid by delta-6 desaturase. Sapienic acid attracts attention as novel marker of cancer cell plasticity. Here, we analyzed fatty acids including the n-10 fatty acid contents, and compared for the first time cell membranes and the corresponding extracellular vesicles (EV) of two human prostatic adenocarcinoma cell lines of different aggressiveness (PC3 and LNCaP). The n-10 components were 9-13% of the total fatty acids in both cancer cell lines and EVs, with total MUFA levels significantly higher in EVs of the most aggressive cell type (PC3). High sapienic/palmitoleic ratios indicated the preference for delta-6 vs. delta-9 desaturase enzymatic activity in these cell lines. The expressions analysis of enzymes involved in desaturation and elongation by qRT-PCR showed a higher desaturase activity in PC3 and a higher elongase activity toward polyunsaturated fatty acids than toward saturated fatty acids, compared to LNCaP cells. Our results improve the present knowledge in cancer fatty acid metabolism and lipid phenotypes, highlighting EV lipidomics to monitor positional fatty acid isomer profiles and MUFA levels in cancer.

Wounding is a serious environmental stress in plants. Oxylipins such as jasmonic acid play an important role in defense against wounding. Mechanisms to adapt to wounding have been investigated in vascular plants; however, those mechanisms in nonvascular plants remain elusive. To examine the response to wounding in Physcomitrella patens, a model moss, a proteomic analysis of wounded P. patens was conducted. Proteomic analysis showed that wounding increased the abundance of proteins related to protein synthesis, amino acid metabolism, protein folding, photosystem, glycolysis, and energy synthesis. 12-Oxo-phytodienoic acid (OPDA) was induced by wounding and inhibited growth. Therefore, OPDA is considered a signaling molecule in this plant. Proteomic analysis of a P. patens mutant in which the PpAOS1 and PpAOS2 genes, which are involved in OPDA biosynthesis, are disrupted showed accumulation of proteins involved in protein synthesis in response to wounding in a similar way to the wild-type plant. In contrast, the fold-changes of the proteins in the wild-type plant were significantly different from those in the aos mutant. This study suggests that PpAOS gene expression enhances photosynthesis and effective energy utilization in response to wounding in P. patens.

Recently we have seen a relaxation of the historic restrictions on the use and subsequent research on the Cannabis plants, generally classified as Cannabis sativa and Cannabis indica. What research has been performed to date has centered on chemical analysis of plant flower products, namely cannabinoids and various terpenes that directly contribute to phenotypic characteristics of the female flowers. In addition, we have seen many groups recently completing genetic profiles of various plants of commercial value. To date, no comprehensive attempt has been made to profile the proteomes of these plants. We report herein our progress on constructing a comprehensive draft map of the Cannabis proteome. To date we have identified over 17,000 potential protein sequences. Unfortunately, no annotated genome of Cannabis plants currently exists. We present a method by which “next generation” DNA sequencing output and shotgun proteomics data can be combined to produce annotated FASTA files, bypassing the need for annotated genetic information altogether in traditional proteomics workflows. The resulting material represents the first comprehensive annotated protein FASTA for any Cannabis plant. Using this annotated database as reference we can refine our protein identifications, resulting in the confident identification of 13,000 proteins with putative function. Furthermore, we demonstrate that post-translational modifications play an important role in the proteomes of Cannabis flower, particularly lysine acetylation and protein glycosylation. To facilitate the evolution of analytical investigations into these plant materials, we have created a portal to host resources we have developed from proteomic and metabolomic analysis of Cannabis plant material as well as our results integrating these resources. All data for this project is available to view or download at www.CannabisDraftMap.Org

Single molecule de novo protein sequencing based on the 'superspecificity' of amino-acyl tRNA synthetases (aaRS) is proposed. An unfolded protein molecule is threaded through a nanopore in an electrolytic cell (e-cell) to expose the terminal residue in the e-cell's trans chamber. After the residue is cleaved with a peptidase, a set of tRNAs, their aaRSs, and ATP are added to trans. An aaRS charges a cognate tRNA molecule with the residue. The charged tRNA (along with the other reactants) is transferred to an extended e-cell with N (20 ≤ N ≤ 61) pores in N individual cis chambers and a single trans chamber. Each pore holds an RNA molecule ending in a unique codon that is exposed in trans. The charged tRNA's anticodon base-pairs with the terminal codon of an RNA. If tRNAs and residues are fluorescently tagged with two different colors, the residue can be identified from the observed position of the color pair. As charging is 'superspecific' identification is unambiguous. The protein molecule in the first e-cell is advanced by one residue and the process repeated. In this approach there is no need for precise pore current or optical intensity measurements. Potential implementation issues are discussed. Several variations, including one in which the terminal residue is cleaved after charging, are also considered.

Molecular visualisation is fundamental in the current scientific literature, textbooks and dissemination materials, forming an essential support for presenting results, reasoning on and formulating hypotheses related to molecular structure. Visual exploration has become easily accessible on a broad variety of platforms thanks to advanced software tools that render a great service to the scientific community. These tools are often developed across disciplines bridging computer science, biology and chemistry. Here we first describe a few Swiss Army knives geared towards protein visualisation for everyday use with an existing large user base, then focus on more specialised tools for peculiar needs that are not yet as broadly known. Our selection is by no means exhaustive, but reflects a diverse snapshot of scenarios that we consider informative for the reader. We end with an account of future trends and perspectives.

The regulation of multiple bacterial phenotypes was found to depend on different cyclic di-nucleotides (CDNs) that constitute intracellular signalling second messenger systems. Most notably, c-di-GMP, along with proteins related to its synthesis, sensing and degradation, was identified as playing a central role in the switching from biofilm to planktonic modes of growth. Recently, this world has been under expansion, with the discoveries of other signalling CNDs in bacteria (c-di-AMP and cGAMP) and also in eukaryotes, novel protein and RNA receptors of CDNs, and the numerous functions related to these molecules. In this work, we comprehensively review and analyse the structural biology data about the systems that bacteria use to synthesise and recognise CDNs, detailing their interactions at molecular level with their products/ligands. Additional interesting observations were made, including that different receptor types can bind CDNs in similar conformations and that, based on genomic data, different CDN second messenger systems may coexist in many organisms. The large amount of sequence and structural data available allows a broad view of the importance of CDNs in bacteria, but how cells coordinate these molecules to ensure adaptation to changing environmental conditions is still open for much further exploration.

The involvement of the angiotensin II type 1 receptor in the Frank-Starling Law of the Heart, where the various activations are very limited, allows simple analysis of the kinase systems involved and thence extrapolation of the mechanism to that of angiotensin control of activation of cardiac and skeletal muscle contraction. The involvement of phosphorylation of the myosin light chain in the control of contraction is accepted but not fully understood. The involvement of troponin-I phosphorylation is also indicated but of unknown mechanism. There is no known signal for activation of myosin light chain kinase or Protein Kinase C-βII other than Ca²⁺/calmodulin but the former is constitutively active and thus has to be under control of a regulated inhibitor, the latter kinase may also be the same. Ca²⁺/calmodulin is not activated in Frank-Starling, i.e. there are no diastolic or systolic [Ca²⁺] changes. I suggest here that that the regulated inhibition is by myosin light chain phosphatase and/or β-arrestin. Angiotensin activation is by translocation of the β-arrestin from the sarcoplasm to the PM thus reducing its kinase inhibition action in the sarcoplasm, this reduced inhibition has been wrongly attributed to a mythical downstream agonist property of β-arrestin.

SMAD ubiquitination regulatory factor 1 (Smurf1) is a Nedd4 family E3 ubiquitin ligase that regulates cell motility, polarity and TGFβ signaling. Smurf1 contains an N-terminal protein kinase C conserved 2 (C2) domain that targets cell membranes and is required for interactions with membrane-localized substrates such as RhoA. Here we investigated the lipid-binding mechanism of Smurf1 C2, revealing a general affinity for anionic membranes in addition to a selective affinity for phosphoinositides (PIPs). We found that Smurf1 C2 localizes not only to the plasma membrane but also to negatively charged intracellular sites, acting as an anionic charge sensor and selective PIP-binding domain. Site-directed mutagenesis combined with docking/molecular dynamics simulations revealed that the Smurf1 C2 domain loop region primarily interacts with PIPs and cell membranes, as opposed to the β-surface cationic patch employed by other C2 domains. By depleting PIPs from the inner leaflet of the plasma membrane, we found that PIP binding is necessary for plasma membrane localization. Finally, we used a Smurf1 cellular ubiquitination assay to show that the amount of ubiquitin at the plasma membrane interface depends on the lipid-binding properties of Smurf1. This study shows the mechanism by which Smurf1 C2 targets membrane-based substrates and reveals a novel interaction based on PI(4,5)P2 and PIP3 selectivity.

Translation is spatiotemporally regulated and ER-associated mRNAs are generally in efficient translation. It is unclear whether the ER-associated mRNAs are deadenylated or degraded on the ER surface in situ or in the cytosol. Here, we showed that ER possessed active deadenylases, particularly the poly(A)-specific ribonuclease (PARN), in common cell lines and mouse tissues. Consistently, purified recombinant PARN exhibited a strong ability to insert into the Langmuir monolayer and liposome. ER-anchored PARN was found to be able to reshape the poly(A) length profile of the ER-associated RNAs by suppressing long poly(A) tails without significantly influencing the cytosolic RNAs. The shortening of long poly(A) tails did not affect global translation efficiency, suggesting that the non-specific action of PARN towards long poly(A) tails was beyond the scope of translation regulation on the ER surface. Transcriptome sequencing analysis indicated that the ER-anchored PARN trigged the degradation of a small subset of ER-enriched transcripts. The ER-anchored PARN modulated the translation of its targets by redistributing ribosomes to heavy polysomes, suggesting that PARN may play a role in dynamic ribosome reallocation. During DNA damage response, MK2 phosphorylated PARN-Ser557 to modulate PARN translocation from the ER to cytosol. By promoting the decay of ER-associated MDM2 transcripts with low ribosome occupancy, the ER-anchored PARN modulated DNA damage response and thereby cell viability. These findings revealed that a highly regulated communication between mRNA degradation rate and translation efficiency is present on the ER surface in situ and that PARN may contribute to this communication by modulating the dynamic ribosome reallocation between transcripts with low and high ribosome occupancies.

The involvement of the angiotensin II type 1 receptor in the Frank-Starling Law of the Heart, where the various activations are very limited, allows simple analysis of the kinase systems involved and thence extrapolation of the mechanism to that of angiotensin control of activation of cardiac and skeletal muscle contraction. The involvement of phosphorylation of the myosin light chain in the control of contraction is accepted but not fully understood. The involvement of troponin-I phosphorylation is also indicated but of unknown mechanism. There is no known signal for activation of myosin light chain kinase or Protein Kinase C-βII other than Ca²⁺/calmodulin but the former is constitutively active and thus has to be under control of a regulated inhibitor the latter kinase may also be the same. Ca²⁺/calmodulin is not activated in Frank-Starling, i.e. there are no diastolic or systolic [Ca²⁺] changes. I suggest here that that the regulated inhibition is by myosin light chain phosphatase and/or β-arrestin. Angiotensin activation is by translocation of the β-arrestin from the sarcoplasm to the PM thus reducing its inhibition in the sarcoplasm, this reduced inhibition has been wrongly attributed to a mythical downstream agonist property of β-arrestin.

Ubiquitin-specific protease 14 (USP14) is a member of the Deubiquitinating enzymes (DUBs) involved in disrupting the regulation of the ubiquitin-proteasome system, responsible for the degradation of impaired and misfolded proteins which is an essential mechanism in eukaryotic cells. The involvement of USP14 in cancer progression and neurodegenerative disorders has been reported. Thereof USP14 is a prime therapeutic target; hence, designing efficacious inhibitors of USP14 is central in curbing these conditions. Herein, we relied on structural bioinformatics methods incorporating molecular docking, Molecular Mechanics Generalized Born Surface Area (MM-GBSA), Molecular dynamics simulation (MD simulation) and ADME to identify potential allosteric USP14 inhibitors. A library of over 733 compounds from the PubChem repository with >90% match to the IU1 chemical structure was screened in a multi-step framework to attain prospective drug-like inhibitors. Two potential lead compounds (CID 43013232 and CID 112370349) were shown to record better binding affinity compared to IU1, but with subtle difference to IU1-47, a 10-fold potent compound when compared to IU1. The stability of the lead molecules complexed with USP14 was studied via MD simulations. The molecules were found to be stable within the binding site throughout the 50ns simulation time. Moreover, the protein-ligand interactions across the simulation run time suggest Phe331, Tyr476, and Gln197 as crucial residues for USP14 inhibition. Furthermore, in silico pharmacological evaluation revealed the lead compounds as pharmacological sound molecules. Overall, the methods deployed in this study revealed two novel candidates that may show selective inhibitory activity against USP14, which could be exploited to produce potent and harmless USP14 inhibitors.

There has recently been interest in the development of small-molecule inhibitors of the oligomerization of Bacillus anthracis protective antigen for therapeutic use. Some of the proposed lead compounds have, however, unfavorable solubility in aqueous medium, which prevents their clinical use. In this computational work, we have designed several hundreds of derivatives with progressively higher hydrosolubility and tested their ability to dock the relevant binding cavity. The highest-ranking docking hits were then subjected to 125 ns-long simulations to ascertain the stability of the binding modes. Several of the potential candidates performed quite disappointingly , but two molecules showed very stable binding modes throughout the complete simulations. Besides the identification of these two promising leads, these molecular dynamics simulations allowed the discovery of several insights that shall prove useful in the further improvement of these candidate towards higher potency and stability.

The concentration of all-trans-retinoic acid, the bioactive derivative of vitamin A, is critically important for the optimal performance of numerous physiological processes. Either too little or too much of retinoic acid in developing or adult tissues is equally harmful. All-trans-retinoic acid is produced by the irreversible oxidation of all-trans-retinaldehyde. Thus, the concentration of retinaldehyde as the immediate precursor of retinoic acid has to be tightly controlled. However, the enzymes that produce all-trans-retinaldehyde for retinoic acid biosynthesis and the mechanisms responsible for the control of retinaldehyde levels have not yet been fully defined. The goal of this review is to summarize the current state of knowledge regarding the identities of physiologically relevant retinol dehydrogenases, their enzymatic properties and tissue distribution, and to discuss potential mechanisms for the regulation of the flux from retinol to retinaldehyde.

Corneal ulcer (CU) is an ophthalmopathy characterized by depression of the corneal surface with at least one stromal loss. CU is common in canine and feline species and is usually caused, among others, by trauma, infections, toxic contamination and endocrine disorders. They usually result from an increased inflammatory response and are associated with some clinical signs such as blepharospasm, photophobia, epiphora, pain and loss of corneal transparency. Despite advances in conventional and pharmacological therapy, in many cases indolent and recurrent ulcer treatments still lead to loss of visual acuity of the animal. This paper aims to report the effect of topical application of canine adipose tissue-derived mesenchymal stem cell (cATMSCs) as treatment of recurrent CU in a Poodle dog breed that showed clear difficulty in the healing process associated with diabetes. The animal was submitted to two applications of cATMSCs and showed improvement in the blepharospasm, conjunctival hyperemia, mucopurulent ocular secretion, photophobia, corneal opacity, chemosis, pigmentation, neovascularization, and pain parameters. Besides, Fluorescein test, Schirmer test and ocular fundus exam also showed improvement in their values concomitantly with lesion resolution. Due this, we showed that cATMSC therapy contribute to the regeneration of corneal tissue in CU and may contribute to the treatment to others ophthalmopathies.

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.

A method for sequencing a protein from a codon sequence is proposed. An unfolded protein molecule is threaded through a nano-sized pore in an electrolytic cell carboxyl end first and held with a voltage such that only the first residue is exposed in the trans chamber of the cell. A tRNA molecule in trans with matching anticodon for the residue binds itself to the latter in the presence of suitable catalysts. It is then cleaved and transferred to an extended electrolytic cell with N pores, 40 ≤ N ≤ 61, in N individual cis chambers and a single trans chamber. Each pore holds an RNA molecule ending in a unique codon that is held exposed in the trans chamber. In the presence of suitable catalysts the anticodon in the transferred tRNA binds with the codon of a matching RNA molecule. By reversing the voltages in the extended cell every RNA molecule except the one to which the transferred tRNA is bound retracts into its cis chamber, this identifies the residue unambiguously. The detected residue in the first cell is cleaved and the process repeated. Unlike in other nanopore-based methods, it suffices to detect the occurrence of a current blockade without having to measure the pore current precisely. A simplified but more time-consuming version that uses only the first cell is also described. In either case no a priori information about the protein is needed so de novo sequencing is possible. A feasibility analysis of the proposed scheme is presented.

Twelve overnight fasted, healthy, male volunteers received on separate days a test breakfast consisting of (A) 100 g fresh white bread (providing 50 g starch) and 250 mL drinking water, (B) the same bread with a supplement of 10 g alpha-cyclodextrin dissolved in the drinking water (250 mL), and (C) 250 mL drinking water containing 25 g alpha-cyclodextrin. Capillary and venous blood samples were collected before breakfast and at regular intervals for a period of 3 hours thereafter. Plasma glucose was determined in capillary blood and plasma insulin in venous blood samples. Breakfast (A) let to the expected rise in blood glucose and insulin concentrations. Breakfast (C) did not produce a significant glycemic and insulinemic response, demonstrating that alpha-cyclodextrin is not hydrolyzed to glucose in the human digestive tract. Mild intestinal symptoms after the ingestion of alpha-cyclodextrin were reported by 4 subjects. The postprandial rises of plasma glucose and insulin were significantly smaller after breakfast (B) than (A). Under the conditions of this study, alpha-cyclodextrin reduced the glycemic and insulinemic index of white bread by 57 and 55 %, respectively. The postprandial time profile of plasma glucose and insulin suggests that, in an initial phase, the digestion of starch is inhibited by alpha-cyclodextrin almost completely. Yet, despite the delayed and reduced digestion of starch, the intake of breakfast (B) was not associated with flatulence or any other gastrointestinal symptoms.

Despite all efforts, the treatment of breast cancer (BC) cannot be considered to be a success story. The advances in surgery, chemotherapy and radiotherapy have not been sufficient. Indeed, the accumulated experience clearly indicates that new perspectives and non-main stream approaches are needed to better characterize the etiopathogenesis and treatment of this disease. This contibution deals with how the new pH-centric anticancer paradigm plays a fundamental role in reaching a more integral understanding of the etiology, etiopathogenesis and treatment of this multifactorial disease. For the first time the armamentarium available for the treatment of the different types and phases of BC is approached here from a Unitarian perspective based upon the hydrogen ion dynamics of cancer. The wide-ranged pH-related molecular, biochemical and metabolic model is able to embrace most the fields and subfields of breast cancer pathology. This single and integrated approach allows to advance a unidirectional program to treatment. Further efforts in this line are likely to first improve the therapeutics of each subtype of this tumor, then every phase of the disease and finally every individual patient.

A method for indirectly sequencing a protein from a codon sequence is proposed. An unfolded protein molecule is threaded through a nano-sized pore in an electrolytic cell carboxyl end first and held with a voltage such that only the first residue is exposed in the trans chamber of the cell. A tRNA molecule in trans with matching anticodon for the residue binds itself to the latter in the presence of suitable catalysts. It is then cleaved and transferred to an extended electrolytic cell with N pores, 40 ≤ N ≤ 61, in N individual cis chambers and a single trans chamber. Each of the N pores holds an RNA molecule ending in a unique codon that is held exposed in the trans chamber. In the presence of suitable catalysts the anticodon in the transferred tRNA binds with the codon of a matching RNA molecule. By reversing the voltages in the extended electrolytic cell every RNA molecule except the one to which the transferred tRNA is bound recedes into its cis chamber, this identifies the residue unambiguously. The detected residue in the first cell is cleaved and the process repeated. A feasibility analysis is presented for the proposed scheme. The method is suitable for de novo sequencing as no a priori information about the protein is needed.

Forest healing effects are increasingly valued for their contribution to human psychological and physiological health, motivating further advances aimed at improving the knowledge of the relevant forest resources. Biogenic volatile organic compounds, emitted by the plants and accumulating in the forest atmosphere, are essential contributors to the forest healing effects, and represent the focus of this study. Using a photoionization detector, we investigated the high frequency variability, in time and space, of the concentration of total volatile organic compounds, on a hilly site, as well as along forest paths and long hiking trails on Italian northern Apennines. The scale of concentration variability was found to be comparable to absolute concentration levels, within time scales of less than one hour, and spatial scales of several hundred meters. During daylight hours, the concentration peaked from noon to early afternoon, followed by early morning, with lowest levels in late afternoon. Based on a conceptual model, these results were related to meteorological variables, including the atmospheric vertical stability profile. Moreover, preliminary evidence pointed to higher concentration of volatile organic compounds in forests dominated by conifer trees, in comparison with pure beech forests.

Directed evolution methods are becoming increasingly popular, as they are extremely powerful toward developing new biomolecules with altered/novel activities, e.g., proteins with new catalytic functions or substrate specificities, and nucleic acids that recognize an intended target. Especially useful are systems that have incorporated continuous evolution, where the protein to be evolved undergoes continuous mutagenesis to evolve a desired trait with little to no input from the researcher once the system is started. However, continuous evolution methods can be challenging to implement in the lab and daunting for researchers to invest time and resources. Our intent is to provide basic information and helpful suggestions that we have gained from our experience with bacterial phage-assisted continuous evolution (PACE). Specifically, we review factors to consider before adopting PACE for a given evolution scheme, different types of selection circuits that can be utilized with particular focus on the PACE-B1H selection system, what optimization of a PACE selection circuit may look like using directed evolution of ME47 as a case study, and additional techniques that may be incorporated into a PACE experiment. With this information, researchers will be better equipped to determine if PACE is a valid strategy to use to evolve their proteins and how to set up a valid selection circuit.

Mammalian cells maintain the complex glycerophospholipid (GPL) class compositions of their various membranes within close limits because this is essential to their well-being or viability. Surprisingly, however it is still not understood how those compositions are maintained except that GPL synthesis and degradation closely coordinated. Here, we hypothesize that abrupt changes in the chemical activity of the individual GPL classes coordinate the synthesis and degradation, as well other homeostatic processes. A previously proposed model proposed that in cellular membranes only a limited number of “allowed” or optimal GPL glass compositions exist because they are energetically more favorable than the other compositions, i.e. they represent local free energy minima (Somerharju et al. 2009). This model, however, could not satisfactorily explain how the optimal compositions are sensed by the key homeostatic enzymes i.e., the rate-limiting synthetizing enzymes and the degrading enzymes (i.e., homeostatic phospholipases). We now propose that when the mole fraction of a GPL class exceeds an optimal one, its chemical activity abruptly increases, which (i) increases its propensity to efflux from the membrane thus making it susceptible for hydrolysis by homeostatic phospholipases, (ii) increases its potency to inhibit its own biosynthesis via a feedback mechanism, (iii) enhances its conversion to another GPL class via a novel process termed “head group remodeling” or (iv) enhances its translocation to other subcellular membranes. Accordingly, abrupt changes in the chemical activity of the individual GPL classes is proposed to regulate and coordinate those four processes maintaining GPL class homeostasis in mammalian cells.

Background: Catalase is a vital antioxidant enzyme that dismutates H₂O₂ into water and molecular oxygen. Many protocols have been developed to measure catalase enzyme activity. Spectrophotometric methods are the most common assays that used to assess catalase enzyme activity. Methods: Because the rate-limiting step during catalase enzyme activity depends upon the dissociation of hydrogen peroxide, the developed assay measures the reaction between a hydroquinone/anilinium sulfate/ammonium molybdate reagent and Unreacted Hydrogen Peroxide, which results in the production of a purple, disubstituted quinone compound with a maximum absorbance value at 550 nm. Results: To clarify the precision of the developed method, the coefficients of variation were determined to be 2.6% and 4.7% for within run measurements and between run measurements, respectively. This method returned results that correlated well (r = 0.9982) with the results returned using the peroxovanadate method to assess catalase enzyme activity. Additionally, we examined the use of the newly developed hydroquinone assay to measure catalase enzyme activity in liver and bacterial homogenate samples. Conclusion: These results demonstrated that this assay can be used for scientific research and routine health applications because it is inexpensive, simple, accurate, and rapid. This method is suitable for use in clinical pathology laboratories because it is simple and produces precise and reproducible results.

To determine the mode of action of the effects of phytoalexins in soybeans, we analyzed enzyme inhibition kinetics using Michaelis–Menten plots and the Lineweaver–Burk plots. The results showed that glyceollin showed competitive inhibition, genistein showed noncompetitive, daidzein was uncompetitive, and luteolin showed a mixed mode of action. The Ki values were determined using a Dixon plot as: glyceollin, 18.99; genistein, 15.42; luteolin, 16.81; and daidzein, 9.99 μM, respectively. Furthermore, potential synergistic effects between glyceollin and the three designated polyphenols were investigated. A combination of glyceollin and luteolin (the ratio of 3:7 of glyceollin and luteolin) had synergistic effects on α-glucosidase inhibition according to combination index (CI)-isobologram equation. Collectively, these results showed that a combination of glyceollin and luteolin has the potential to inhibit α-glucosidase activity via a synergistic mode of action.

This study focused on hydrolysis of cosmetic fillers hyaluronic acid (HA) and kinetics of the HA hydrolysis using the homogenate of the red king crab hepatopancreas. Turbidimetric analysis of the reaction mixture revealed a bell-shaped time dependence of aggregation formation. It was shown that the obtained homogenate has the similar activity to the commercially available hyaluronidase. The atomic force microscopy (AFM) examination found that the HA fillers were represented by spherical-like structures. These structures were destroyed under the action of the homogenate of the red king crab hepatopancreas. NMR of the reaction mixture showed that HA degradation lasts for some days, but a maximum rate of the reaction is detected in the first hours of incubation. The preparation with hyaluronidase activity obtained from the red king crab hepatopancreas could be used as potentially safe product for treating filler complications.

In an attempt to correct misunderstandings this article brings together the observations on Calcium, Myosin Binding Protein-C and Hypertrophic Cardiomyopathy in the basic function of cardiac muscle. A finding of many years ago is reiterated in a novel enzyme kinetic format with defined rate limiting step which makes CaATP the apparent substrate of the actomyosin cross-bridge. The relationship of these kinetics to recent observations on disruption of myosin binding protein-C is described along with how this bears on the understanding of the related cardiomyopathies.

Significance: Electrophile signaling is coming into focus as a bona fide cell signaling mechanism. The electrophilic regulation occurs typically through a sensing event (i.e., labeling of a protein) and a signaling event (the labeling event having an effect of the proteins activity, association, etc.). Recent advances: Herein, we focus on the first step of this process, electrophile sensing. Electrophile sensing is typically a deceptively simple reaction between the thiol of a protein-cysteine, of which there are around 200,000 in the human proteome, and a Michael acceptor, of which there are numerous flavors, including enals and enones. Recent data overall paint a picture that despite being a simple chemical reaction, electrophile sensing is a discerning process, showing labeling preferences that are often not in line with reactivity of the electrophile. Critical issues: With a view to trying to decide what brings about highly electrophile-reactive protein-cysteines, and how reactive these sensors may be, we discuss aspects of the thermodynamics and kinetics of covalent/non-covalent binding. Data made available by several laboratories indicate that it is likely that specific proteins exhibit highly stereo- and chemoselective electrophile sensing, which we take as good evidence for recognition between the electrophile and the protein prior to forming a covalent bond. Future directions: We propose experiments that could help us gain a better and more quantitative understanding of the mechanisms through which sensing comes about. We further extoll the importance of performing more detailed experiments on labeling and trying to standardize the way we assess protein-specific electrophile sensing.We propose experiments that could help us gain a better and more quantitative understanding of the mechanisms through which sensing comes about. We further extoll the importance of performing more detailed experiments on labeling and trying to standardize the way we assess protein-specific electrophile sensing.

This study aimed to determine the Gas Chrommatography (GC)-Mass Spectrometry (MS) profiles and insecticidal activity of essential oils (EOs) from Thymus vulgaris (Thyme) and Cymbopogon citratus (Lemongrass) against the invasive and devastating pest, Tuta absoluta (T. absoluta) through contact and fumigation routes. We found out that thyme oil was predominantly constituted of Thymol (22.16%), α-Pinene (15.35%) and p-Cymene (13.54%) whilst Neral (21.41%), Geranial (21.36%) and β-Myrcene (9.74%) were the major constituents of lemongrass oil. Lemongrass oil exhibited higher insecticidal efficiency irrespective of application mode with 50% lethal dose (LD50) values of 35.8 and 72.2 µL.L-1air on contact and fumigation routes, respectively. Lemongrass oil also lengthened pupal duration at all tested doses irrespective of application routes. The overall responses of Lemongrass oil surpassed that of the reference insecticide (Lynx®: Lambda-cyhalothrine; Acetamipride). Thus, the recorded data clearly showed the acute and long-term insecticidal effects of the studied EOs, though a greenhouse and open field trials are required prior to the validation of this approach as remediation measure for Integrated Pest Management (IPM) for tomato borer control in Cameroon and elsewhere.

Autotaxin (ATX) is a secreted lysophospholipase D, catalysing the conversion of lysophosphatidylcholine (LPC) to bioactive lysophosphatidic acid (LPA). LPA acts through two families of G protein-coupled receptors (GPCRs) controlling key cellular responses, and is implicated in many physiological processes and pathologies. ATX has therefore been established as an important drug target in the pharmaceutical industry. Structural and biochemical studies of ATX have shown that it has a bimetallic nucleophilic catalytic site, a substrate-binding (orthosteric) hydrophobic pocket that accommodates the lipid alkyl chain, and an allosteric tunnel that can accommodate various steroids and LPA. Here we first review what is known about ATX-mediated catalysis, crucially in light of allosteric regulation. We then present the known ATX catalysis-independent functions, including binding to cell-surface integrins and proteoglycans. In light of these data we then discuss the four types of ATX inhibitors, as classified depending on their binding to the orthosteric and/or the allosteric site. Finally, we analyse the binding mode of known members of all four types and discuss how mechanistic differences might differentially modulate the activity of the ATX-LPA signalling axis, and clinical applications including cancer.

Adlay (Coix lachryma-jobi L. var. ma-yuen Stapf) contains various phytonutrients for treating many diseases in Asia. To investigate whether orally administered of adlay bran oil (ABO) can cause drug interactions, the effects of ABO on the pharmacokinetics of five cytochrome P450 (CYP) probe drugs were evaluated. Rats were given a single oral dose (2.5 mL/kg BW) of ABO 1 h before administration of a drug cocktail either orally or intravenously, and blood was collected at various time points. A single oral dose of ABO administration did not affect the pharmacokinetics of five probe drugs when given as a drug cocktail intravenously. However, ABO increased plasma theophylline, dextromethorphan, and diltiazem when co-administered an oral drug cocktail. After 7-days of feeding with an ABO-containing diet, plasma concentrations of theophylline and chlorzoxazone were increased after oral administration of drug cocktail. The major CYP enzyme activities in liver and intestinal were not affected by ABO treatment. Results from this study indicate that a single oral dose or short-term administration of ABO may increase plasma drug concentrations when ABO is given concomitantly with drugs. ABO is likely to enhance intestinal drug absorption. Therefore, caution is needed to avoid food-drug interactions between ABO and co-administered drugs.

Environmental interventions and ecological adaptations harbor millions of valued substances and metabolites in plants which can be employed and commercialized for human benefits. Present study encompasses the untapped potential of pine needles of Indo-Himalayan region for the production of different metabolites and their pharmacological significance in terms of antioxidant and antimicrobial activity. Total phenolic and flavonoid content from the needles of ten pine species was quantified using three different solvent systems. Results revealed that out of 10 different selected Pinus species, Pinus taeda showed highest concentration of total phenolics, soluble-F phenolics and flavonoids content (approx.147.02 mg/g, 141.08 mg/g and 21.91mg/g respectively) as compared to other species. On the other hand P. greggii showed highest Bound-W phenolic content (approx.3.62mg/g). Among all the selected plant species, the needles of P.echinata exhibited the highest and P.thunbergii had the lowest ratio of total flavonoids to total phenolics. Most of these compounds were found to have effective antioxidant activities as well as antimicrobial activity, as estimated by oxygen radical absorbance capacity (ORAC) and disk diffusion test respectively.

The kinetics of the hydrolysis of hyaluronic acid (HA) of cosmetic fillers using thehomogenate of the red king crab hepatopancreas was studied for the first time. Turbidimetricanalysis of the reaction mixture revealed a bell–shaped time dependence of aggregation formation. The HA fillers were examined by atomic force microscopy (AFM) and it was found that they wererepresented by spherical–like structures. These structures were disrupted under the action of thehomogenate of the red king crab hepatopancreas. It was shown that the prepared homogenate hasthe activity which is similar to that observed in the commercially available hyaluronidase products.The preparation with hyaluronidase activity obtained from the red king crab hepatopancreas couldbe used as potentially safe product for treating filler complications.

In addition to the canonical loss-of-function mutations, mutations in proteins may additionally result in gain-of-function through the binary activation of cryptic ‘structural capacitance elements’. Our previous bioinformatic analysis allowed us to propose a new mechanism of protein evolution - structural capacitance – that arises via the generation of new elements of microstructure upon mutations that cause a disorder-to-order (DO) transition in previously disordered regions of proteins. Here we propose that the DO transition is a necessary follow-on from expected early codon-anticodon and tRNA acceptor stem-amino acid usage, via the accumulation of structural capacitance elements - reservoirs of disorder in proteins. We develop this argument further to posit that structural capacitance is an inherent consequence of the evolution of the genetic code.

The limited information available on the structure of complexes involving transcription factors and cognate DNA response elements represents a major obstacle in the quest to understand their mechanism of action at the molecular level. We implemented a concerted structural proteomics approach, which combined hydrogen-deuterium exchange (HDX), quantitative protein-protein and protein-nucleic acid cross-linking (XL), and homology analysis, to model the structure of the complex between the full-length DNA binding domain (DBD) of FOXO4 and its DNA binding element (DBE). The results confirmed that FOXO4-DBD assumes the characteristic forkhead topology shared by these types of transcription factors, but its binding mode differs significantly from those of other members of the family. The results showed that the binding interaction stabilized regions that were rather flexible and disordered in the unbound form. Surprisingly, the conformational effects were not limited only to the interface between bound components but extended also to distal regions that may be essential to recruiting additional factors to the transcription machinery. In addition to providing valuable new insights into the binding mechanism, this project provided an excellent evaluation of the merits of structural proteomics approaches in the investigation of systems that arematerial not directly amenable to traditional high-resolution techniques.

Research on healthy ageing shows that lifespan reductions are often caused by mitochondrial dysfunction. Thus, it is very interesting that the deletion of mitochondrial matrix peptidase LonP1 was observed to abolish embryogenesis, while deletion of the mitochondrial matrix peptidase ClpP prolonged survival. To unveil the targets of each enzyme, we documented the global proteome of LonP1^+/- mouse embryonal fibroblasts (MEF), for comparison with ClpP^-/- depletion. Proteomic profiles of LonP1^+/- MEF generated by label-free mass spectrometry were further processed with the STRING webserver Heidelberg for protein interactions. ClpP was previously reported to degrade Eral1 as a chaperone involved in mitoribosome assembly, so ClpP deficiency triggers accumulation of mitoribosomal subunits and inefficient translation. LonP1^+/- MEF also showed Eral1 accumulation, but no systematic effect on mitoribosomal subunits. In contrast to ClpP^-/- profiles, several components of the respiratory complex I membrane arm were accumulated, whereas the upregulation of numerous innate immune defense components was similar. Overall, LonP1 as opposed to ClpP appears to have no effect on translational machinery, instead it shows enhanced respiratory dysfunction; this agrees with reports on the human CODAS syndrome caused by LonP1 mutations.

Using a combination of pan proteomic platform associated with systemic biology analyses, we demonstrate that neonatal microglial cells derived from cortex and spinal cord expressed different phenotypes upon the physiological or pathological conditions. They also highlight great variability in protein production on both cellular and exosome levels. Bioinformatics data indicate for the cortical microglia anti-inflammatory and neurogenesis/tumorigenesis characteristics, while for the spinal cord microglia involvement in the inflammatory response. We confirmed these results by performing functional testing including neurite outgrowth assays in DRGs cell line, and glioma proliferation analysis in 3D spheroid cultures. Results from these in vitro assays indicate that the microglia located at different CNS areas reveal differential biological functions. While both microglia sources enhanced growth of DRGs axons, only the spinal microglia significantly attenuated glioma proliferation. Overall these findings are pointing to the fact that the origin of neonatal microglia affects the physio-pathological function, which may address the prevalence of the glioma in the brain in comparison with the spinal cord in adult.

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.

The human proteome is composed from diverse and heterogeneous gene products/proteoforms. Previously, we have been discussing the main technical aspects in developing for inventory of human proteoforms that would be visually attractive, clear, and easy to search (Naryzhny S. J. Proteomics 2018, S1874-3919(18) 30220-3). Here, we present our first draft of the database of proteoforms that is based on this discussion. The database principles and structure are described. The database is called “2DE-pattern” as it contains multiple isoform-centric patterns of proteoforms separated according to 2DE principles.