Benzylsuccinate is generated in anaerobic toluene degradation by radical addition of toluene to fumarate and further degraded to benzoyl-CoA by a beta-oxidation pathway. Using metabolic modules for benzoate transport and activation to benzoyl-CoA and the enzymes of benzylsuccinate beta-oxidation, we established an artificial pathway for benzylsuccinate production in Escherichia coli, which is based of its degradation pathway running in reverse. Benzoate is supplied to the medium, but needs to be converted to benzoyl-CoA by an uptake transporter and a benzoate-CoA ligase or CoA-transferase. In contrast, the second substrate succinate is endogenously produced from glucose under anaerobic conditions, and the constructed pathway includes a succinyl-CoA:benzylsuccinate CoA-transferase that activates it to the CoA-thioester. We present first evidence for the feasibility of this pathway and explore product yields under different growth conditions. Compared to aerobic cultures, the product yield increased more than 1000-fold in anaerobic glucose-fermenting cultures and showed further improvement under fumarate-respiring conditions. An important bottleneck to overcome appears to be product excretion, based on much higher recorded intracellular concentrations of benzylsuccinate, compared to those excreted. While no export system is known for benzylsuccinate, we observed an increased product yield after adding an unspecific mechanosensitive channel to the constructed pathway.

The phenylpropanoid pathway is a major secondary metabolite pathway that helps plants overcome biotic and abiotic stress and produces various by-products that promote human health. Its byproduct, chloroquinic acid (CQA), is a soluble phenolic compound present in many angiosperms. Hy-droxycinnamate-CoA shikimate/quinate transferase(BAHDs superfamily enzyme) is a significant en-zyme that plays a role in accumulating CQA biosynthesis. This study analyzed transcriptome-wide identification of the phenylpropanoid to chloroquinic acid biosynthesis candidate genes in A. spathulifolius flowers and leaves. Transcriptomic analyses of the flowers and leaves showed a differential expression of the PPP and CQA biosynthesis regulated unigenes. An analysis of PPP captive unigenes revealed the following: the major duplication of the key enzyme, PAL, 120 unigenes in leaves and 76 in flowers; the gene encoding C3’H, 169 unigenes in leaves and 140 unigenes in flowers; duplicated unigenes of 4CL, 41 in leaves and 27 in flowers. In addition, C4H unigenes had 12 unigenes in the leaves of A. spathulifolius and four in the flowers. The characterization of the BAHDs superfamily members identified 82 in leaves and 72 in flowers. Among them, phylogenetic analysis showed that five unigenes encoded HQT and three en-coded HCT in A. spathulifolius. The three HQT are common to both leaves and flowers, whereas the two HQT were specialized for leaves. The pattern of HQT synthesis was upregulated in flowers, whereas HCT was expressed strongly in the leaves of A. spathulifolius. Overall, 4CL, C4H, and HQT are expressed strongly in flowers, and caffeic acid and HCT show more expression in leaves. Therefore, CQA biosynthesis occurs in the flowers of A. spathulifolius rather than leaves.

This article presents a method that uses a semi-automatic algorithm for the detection and analysis of microcracks in steel test specimens using binarized image analysis. It obtains the length along the crack path in contrast to previous methods that have obtained linear length, thereby achieving a closer approximation to its actual value. Additionally, this method automatically calculates the COD and COA parameters with measurement deviations in micrometres, thus providing greater precision than previous methods proposed to date.

Proprotein convertase subtilisin/Kexin type 9 (PCSK9) is a proteolytic enzyme that indirectly regulates serum LDL cholesterol by destroying LDL receptors. In clinical studies, the main role of the proprotein convertase subtilisin/Kexin type9 (PCSK9) inhibitor in cholesterol regulation was elucidated. It is produced in the liver but is also present in the kidney and intestine. It prevents HMGCo from synthesizing cholesterol. SREBP-2 is a reductase that is induced by statins. In a dose-dependent manner, increasing SREBP-2 levels enhanced LDL-R and PCSK9 gene expression. At the minimum, two procedures have been developed to overcome the plasma level of PCSK9. This is the LDLR test, polyclonal antibodies, and sentience oligonucleotide. Lower dosage statin treatment with a proprotein convertase subtilisin/Kexin type9 inhibitor will be most efficient in lowering LDL and avoiding statin adverse effects. In multiple long-term trials, statins have been found to reduce cardiovascular mortality by 30% and stroke incidence by 20%. In this way, we conclude the role of PCSK9 in hypercholesterolemia.

Inosine 5'-monophoaphate (IMP) allows animals to sense umami. Intake of IMP in C57/KsJ-db/db (db/db) mice induced lipohyperplasia causing the liver cirrhosis, however, how to that of in normal mammals injure health is still unclear. Thus, we had investigated that intake of IMP in C57BL/6J mice effected its metabolic functions. We found that intake of 255 M/d IMP in C57BL/6J mice for 4 months induced hyperlipidemia and body fat rate raised. In mechanism, the expressions of ACC1 and phosphorylated ACC2 in hepatocytes were increased though IMP promoting phosphorylation of AMPK. The increased ACC1 promoted the conversion of acetyl-CoA into TG. These TG were transported out of hepatocytes to avoid NAFLD, causing a deficiency of acetyl-CoA in liver, and then the increased phosphorylated ACC2 promoted cytoplasm fatty acids into mitochondria to convert into acetyl-CoA though the fatty acids β-oxidation pathway, causing a deficiency of fatty acids. Therefore, liver enhanced the absorption of exogenous fatty acids, which were converted into TG caused lipohyperplasia. Moreover, intake of IMP in normal mice induced complement system weaken in liver causing mild inflammation. Our data not only alerted that humans avoid excessive intake of IMP, but also provided novel insights into the adipose of metabolic dysfunctions.

High production cost is one of the major factors that limit the market growth of polyhydroxyalkanoates (PHAs) as a biopolymer. Improving PHA synthesis performance and utilizing low grade feedstock are two logical strategies for reducing the cost. As an oleaginous yeast, Y. lipolytica has a high carbon flux through acetyl-CoA (main PHB precursor) which makes it a desired cell factory for PHB biosynthesis. Two different metabolic pathways introduced into Y. lipolytica PO1f in this study for enhancing PHB biosynthesis, along with glucose and acetate as co-substrates. Results showed that NBC pathway led to more than 11% PHB accumulation, while there was non-detectable PHB accumulations using ABC pathway. Further modifications were done using peroxisomal compartmentalization and gene-dose overexpression. Final strain showed up to 41% PHB of CDW with a growth rate of 0.227 h-1. Low growth rate was observed using acetate or glucose as sole carbon and energy source. The co-substrate strategy showed compensatory effect on overcoming inhibitory and toxic effects of both substrates at high concentrations. Cultivating the engineered strain in optimal co-substrate condition predicted by RSM led to 83.4 g/L biomass concentration and 31.7 g/L PHB.

Little is known about the effect of statin use in lung cancer development in idiopathic pulmonary fibrosis (IPF). We analyzed the database of the National Health Insurance Service to further investigate the clinical impacts of statin on lung cancer development and overall survival (OS) in IPF patients. The analysis included 9,182 individuals diagnosed with IPF, of which 3,372 (36.7%) were statin users. Compared to statin non-users, the time from diagnosis of IPF to lung cancer development and OS were longer in statin users in IPF patients. In Cox proportional hazard regression models, higher statin compliance, statin use, and being female had an inverse association with lung cancer risk, while older age at diagnosis of IPF and smoking history were associated with higher risk of lung cancer in IPF patients. For OS, statin use, female sex, higher exercise frequency, and diabetes were associated with longer survival. In contrast, older age at diagnosis of IPF and smoking history were associated with shorter OS in IPF patients. These data from a large population indicate that statin had an independent protective association with lung cancer development and mortality in IPF patients.

Advanced cutaneous melanoma is considered the most aggressive type of skin cancer with variable rates of treatment response. Nowadays, there are available some classes of immunotherapy and target therapy for its treatment. Immunotherapy can inhibit tumor growth and its recurrence by triggering host's immune system, while targeted therapy acts inhibiting specific molecules or signaling pathways. However, melanoma response to these treatments are highly heterogenous, and frequently become resistant. Epigenome (DNA/histone modification) contribute to cancer initiation and progression. Epigenetic alterations are divided into four levels of gene expression regulation: DNA methylation, histone modification, chromatin remodeling and non-coding RNAs regulation. The deregulation of lysine methyltransferase enzymes is associated with tumor initiation, invasion, development of metastases, changes in immune microenvironment and drug resistance. The study of lysine histone methyltransferase (KMT) inhibitors is important to cancer epigenetic mechanisms understanding and to biological processes. In addition to immunotherapy and target therapy, the research and development of KMT inhibitors is ongoing. Many studies are exploring the therapeutic implications and possible side effects of these compounds, besides their adjuvant potential to the approved current therapies. Importantly, as with any drug development, safety, efficacy, and specificity are crucial considerations when developing KMT inhibitors for clinical applications.

The glioma-associated oncogene (GLI) family consists of GLI1, GLI2, and GLI3 in mammals, and is the effector in the Hedgehog signaling pathway. This family has important roles in the development and homeostasis of various tissues. To achieve these roles, the GLI family has widespread outputs. GLI activity is therefore strictly regulated at multiple levels, including via post-translational modifications for context-dependent GLI target gene expression. Conversely, dysregulated GLI activation has strong links with a variety of cancers. The protein arginine methyl transferase (PRMT) family is also associated with embryogenesis, homeostasis, and cancer via epigenetic modifications and signal transduction. In the PRMT family, PRMT1, PRMT5, and PRMT7 reportedly regulate GLI1 and GLI2 activity. PRMT1 methylates GLI1 to upregulate its activity and target gene expression. Cytoplasmic PRMT5 methylates GLI1 and is involved in GLI1 protein stabilization. In contrast, nucleic PRMT5 interacts with MENIN to suppress growth arrest-specific protein 1 expression, which assists Hedgehog ligand binding to Patched, indirectly resulting in downregulated GLI1 activity. PRMT7-mediated GLI2 methylation upregulates its activity through the dissociation of GLI2 and Suppressor of Fused. Therefore, PRMT1, PRMT5, and PRMT7 regulate GLI activity at multiple levels, and PRMT-mediated GLI dysregulation may be involved in cancer formation.

Milk fat production is important in the New Zealand (NZ) dairy industry. Elsewhere, an amino acid substitution (K232A) in the enzyme diacylglycerol acyl-CoA acyltransferase (DGAT1) has been reported to explain variation in some milk traits, including variation in milk fatty acid (FA) profile. In this study, associations between K232A, and milk traits including milk FA composition, were studied in wholly pasture-fed NZ Holstein-Friesian × Jersey (HF×J) cross-bred dairy cows. With a high frequency of K variant (61.9%), the KK cows produced more milk fat than the AA cows (5.41 ± 0.04 % vs 4.42 ± 0.05 %). The milk volume, fat concentration and protein concentration of AK cows were between the genotypes, AA and KK. More C16:0, CLA and C18:3 cis-9, 12, 15 FA were found in the milk of Kiwi-cross cows feed in outdoor pasture grazing system., and the influence of DGAT1 K232A, on these FAs from mid- and late lactation stages were significant. The AA cows produced (P < 0.001) more CLA and C18:3 cis-9, 12, 15, but less C16:0 (1.137 ± 0.047, 0.855 ± 0.015 and 35.170 ± 0.355) than the KK cows (0.934 ± 0.025, 0.778 ± 0.009 and 38.010 ± 0.250).

Benign recurrent intrahepatic cholestasis (BRIC) is a rare genetic cause of cholestasis. It is considered as part of inherited intrahepatic cholestasis syndromes, such as progressive familial intrahepatic cholestasis (PFIC), and intrahepatic cholestasis of pregnancy. BRIC is presented in infancy or in early adulthood. It is characterized by exacerbations and remissions of jaundice with accompanying intense itching, lasting from weeks to years throughout lifetime. Normal gamma-glutamyl transferase (GGT) is a characteristic laboratory finding. Contrary to PFIC, which may progress to cirrhosis, BRIC does not progress to chronic liver disease or cirrhosis. However, incessant episodes of cholestasis result in marked reduction in quality of life and distinct mutations increase the risk of hepatobiliary malignancy. In intervals between the exacerbations, the histological findings of centrilobular cholestasis together with the abnormal laboratory parameters return to normal. In this context, liver biopsy might be avoided. In this review, we will focus on the genetic aspects of BRIC, its pathophysiology, clinical presentation, and prognosis of this autosomal recessive genetically determined cholestatic disorder. Moreover, triggering factors as well as treatment options will be further elucidated.

The genus Euonymus (L.) consists of shrubs or woody plants, distributed mainly in the Northern Hemisphere. To date, from Euonymus spp. several hundreds of different secondary metabolites have been isolated and identified. In addition, fatty oil was found in the fruits of some species of these plants, which accumulates not only in the seeds, but also in the arils. This study presents the research of unique long-term (ten-year-old) suspension cell cultures of Euonymus maximoviczianus Prokh. which were obtained from arils of unripe capsules. Suspension cells retain the ability to form oil droplets containing neutral lipids, both in the dark and in the light, the cells are able to synthesize very-long-chain fatty acids (VLCFAs), and can synthesize delphinidin-3-O-hexoside, cyanidin-3-O-hexoside, and peonidin-3-O-hexoside. Here, we research of the fatty acid, VLCFA, and anthocyanin biosynthesis dynamics in the subcultivation, as well as the influence of methyl jasmonate (MeJa) on these processes in the dark- and under-light-grown cell cultures. In the darkness, the formation of VLCFAs is more intense, and the biosynthesis of anthocyanins is significantly activated in the light. MeJA substantially enhances the biosynthesis of anthocyanins in the light and, surprisingly, the formation of VLCFAs in the darknss. In connection with the commonality of the cytosolic pool of malonyl-CoA, which is necessary both for the biosynthesis of VLCFAs and dihydroflavonols (and, ultimately, anthocyanins), the competition of these biosynthetic pathways is discussed.

Coordination of cellular biological processes is regulated in part via metabolic enzymes acting to match cellular metabolism to current conditions. The acetate activating enzyme, acyl-coenzyme A synthetase short-chain family member 2 (Acss2) has long been considered to have a predominantly lipogenic function. More recent evidence suggests that this enzyme has regulatory functions in addition to its role in providing acetyl-CoA for lipid synthesis. We used Acss2 knockout mice (Acss2-/-) to further investigate the roles this enzyme plays in three physiologically distinct organ systems that make extensive use of lipid synthesis and storage including the liver, brain and adipose tissue. We examined the resulting transcriptomic changes resulting from Acss2 deletion and assessed these changes in relation to fatty acid constitution. We find that loss of Acss2 leads to dysregulation of numerous canonical signaling pathways, upstream transcriptional regulatory molecules, cellular processes, and biological functions, which were distinct in the liver, brain and mesenteric adipose tissues. The detected organ-specific transcriptional regulatory patterns reflect the complementary functional roles of these organ systems within the context of systemic physiology. While alterations in transcriptional states were evident, the loss of Acss2 resulted in few changes in fatty acid constitution in all three organ systems. Overall, we demonstrate Acss2 loss institutes organ-specific transcriptional regulatory patterns reflecting the complementary functional roles of these organ systems. Collectively, these findings provide further confirmation that Acss2 regulates key transcription factors and pathways under well-fed, non-stressed conditions, and acts as a transcriptional regulatory enzyme.

This study employs molecular docking experiments to investigate the binding interactions of Hypericin with key proteins implicated in Alzheimer's disease (AD). Hypericin demonstrated a significant binding energy of -10.1 kcal/mol with ABAD (3-hydroxyacyl-CoA dehydrogenase type II). The analysis revealed multiple binding interactions, including hydrogen bonds with arginine 6, serine 7, and glutamic acid 140, as well as hydrophobic bonds involving leucine 11, aspartic acid 142, and aspartic acid 86. These findings illuminate the intricate molecular interactions between Hypericin and ABAD, suggesting its potential as a lead compound for further therapeutic development in the context of Alzheimer's disease. Additionally, a parallel docking investigation was conducted for Beta-secretase 1 (BACE1), a protein pivotal in the generation of beta-amyloid peptides—a hallmark of Alzheimer's disease. Inhibiting BACE1 activity is considered a promising therapeutic strategy for AD. Computational analyses suggest that Hypericin may serve as a potential inhibitor of BACE1. Molecular Docking results reveal a robust binding affinity between Hypericin and BACE1, supported by a significant binding energy score of -10.3 kcal/mol. Notably, both ABAD and BACE1 exhibit high binding affinity with Hypericin. This theoretical exploration provides an initial understanding of the intricate molecular pathways involved in Alzheimer's disease and proposes Hypericin as a potential therapeutic compound for mitigating its progression.

Cross-species investigations on cancer invasiveness are a new approach that identified new biomarkers potentially useful for improving tumor diagnosis and prognosis in clinical medicine and veterinary science. In this study, we combined proteomic analysis of four experimental rat malignant mesothelioma (MM) tumors with analysis of ten patient-derived cell lines to identify common features associated to mitochondrial proteome rewiring. The comparison of significant abundance changes between invasive and non-invasive rat tumors rat tumors gave a list of 433 proteins, including twenty-six proteins reported to be exclusively located in mitochondria. Next, we analyzed the differential expression of genes encoding the mitochondrial proteins of interest in five primary epithelioid and five primary sarcomatoid human MM cell lines, and the most impressive increase was observed in the expression of the long-chain acyl coenzyme A dehydrogenase (ACADL). To evaluate the role of this enzyme in the migration/invasiveness, two epithelioid and two sarcomatoid human MM cell lines derived from patients with the highest and lowest overall survival were studied. Interestingly, sarcomatoid vs epithelioid cell lines were characterized by higher migration and fatty oxidation rates, in agreement with ACADL findings. These results suggest that evaluating mitochondrial proteins in MM specimens might identify tumors with higher invasiveness.

Salidroside, a valuable phenylethanoid glycoside, is obtained from plants belonging to the Rhodiola genus, known for its diverse biological properties. At present, salidroside is still far from large-scale industrial production due to the lower titer and higher process cost. In this study, we constructed a biosynthetic pathway from L-tyrosine to salidroside in Saccharomyces cerevisiae. To increase sali-droside production, we for the first time constructed an in vivo UDP-glucose regeneration system that sucrose synthase GmSUS worked in conjunction with UDP-glucose transferase RrUGT33 to improve the UDP-glucose availability with inexpensive sucrose as the consumed substrate. Finally, the molar conversion rate of L-tyrosine could reach 95%. To further reduce the cost, co-culture with E. coli that synthesized L-tyrosine de novo resulted in an 81% reduction in the final substrate cost per 1 kg of salidroside compared to the initial cost. The final titer of salidroside obtained 3.8 g/L and was the highest titer achieved at the Shake-Flask-Level in the existing reports. And this marked the first successful synthesis of salidroside in an in situ enhanced UDP-glucose system using sucrose. This accomplishment laid a robust groundwork for further investigations into the synthesis of other notable glycosides and natural compounds.

Sporisorium scitamineum, the basidiomycetous fungus causing sugarcane smut that leads to severe losses in sugarcane quantity and quality, undergoes sexual mating to form dikaryotic hyphae capable of invading the host cane. Therefore, suppressing dikaryotic hyphae formation would potentially be an effective way to prevent host infection by the smut fungus, and the following disease symptom development. The phytohormone methyl jasmonate (MeJA) has been showed to induce plant defense against insects and microbial pathogens. In this study, we verified that exogenous addition of MeJA suppressed dikaryotic hyphae formation under in vitro culture condition, and effective in inhibiting maize smut disease symptom caused by Ustilago maydis. We constructed an Escherichia coli strain expressing plant JMT gene, encoding a jasmonic acid carboxyl methyl transferase that catalyzes conversion from jasmonic acid (JA) to MeJA. By GC-MS we confirmed that the transformed E. coli strain, designated as pJMT strain, was able to produce MeJA in the presence of JA and S-adenosylmethionine (SAM; as methyl donor). Furthermore, the pJMT strain was able to suppress S. scitamineum filamentous growth under in vitro culture condition. It waits to further optimize JMT expression under field condition, in order to utilize the pJMT strain as a biocontrol agent (BCA) of sugarcane smut disease. Overall, our study provides a potentially novel method for controlling crop fungal diseases by boosting phytohormone biosynthesis.

This study reviewed aspects of the biology of two members of the glucosinolate family, namely sinigrin and glucoraphanin and their potential biomedical therapeutic and industrial applications. Sinigrin and glucoraphanin are converted by the -sulphoglucosidase myrosinase or the gut microbiota into their bioactive forms, allyl isothiocyanate (AITC) and sulphoraphanin (SFN) which constitute part of a sophisticated defence mechanism plants have developed over several hundred million years of evolution to protect them from parasitic attack from aphids, ticks and nematodes. These compounds display biological activities in a number of mammalian physiological processes and potential biotherapeutic application. Glucosinolates may be useful in bio-fumigation and treatment of biofilms which occur on plant equipment and medical implants formed by problematic pathogenic bacteria such as Pseudomonas aeruginosa. AITC and SFN display similar antibiotic activity as Vancomycin in the treatment of bacteria listed by the World Health Organization as antibiotic-resistant "priority pathogens".AITC and SFN also display bioactivity in cancer chemoprevention through the induction of phase II antioxidant enzymes which inactivate potential carcinogens. The glucosinolates have found application in the prevention of bacterial and fungal spoilage of food substances during processing and in advanced food packaging formats which improve the shelf-life of food products.

The objective of this study was to develop an ultrasonic-assisted procedure for the extraction of total phenolics from Citrus aurantium L. blossoms (CAB) and evaluate the free radical scavenging activity, anti-HMG-CoA reductase activity of total phenolics. In this work, a Box-Behnken design based on the single-factor experiments was used to explore the optimum extraction process. Under the optimum conditions (extraction solvent 70.31% ethanol, extraction temperature 61.94 °C, extraction time 51.73 min and liquid-to-solid ratio 35.63 mL/g), the extraction yield of total phenolics was 95.84 mg gallic acid equivalents (GAE)/g dry matter (DM), which was highly consistent with the theoretical value (96.12 mg GAE/g DM). The total phenolic extract showed excellent free radical scavenging properties against DPPH·, ABTS+·, ·OH and ·O2-, with the IC50 values of 197.007, 83.878, 218.643 and 158.885 μg/mL, respectively, and the extracts also showed good inhibition of HMG-CoA reductase activity, with the IC50 value of 117.165 μg/mL. Total phenolics from CAB could be a potential source of natural free radical scavenger and HMG-CoA reductase inhibitor.

A two-fold integrated research study was conducted; firstly, to understand effects of copper (Cu) and zinc (Zn) on the growth and oxidative stress in Nile tilapia, Oreochromis niloticus; secondly, to study the beneficial effects of the duckweed Lemna minor L. as a heavy metal remover from wastewater. Experiments were conducted in mesocosms with and without duckweed. Tilapia fingerlings were exposed to Cu (0.004 and 0.02 mg/L) and Zn (0.5 and 1.5 mg/L) and fish fed for four weeks. We evaluated the fish growth performance, the hepatic DNA structure using comet assay, the expression of antioxidative genes (superoxide dismutase, SOD; catalase, CAT; glutathione peroxidase, GPx and glutathione-S-transferase, GST) and GPx and GST enzymatic activity. The results showed that Zn exhibited more pronounced toxic effects than Cu. Low dose of Cu did not influence the growth whereas higher doses of Cu and Zn significantly reduced the growth rate of tilapia compared to control, but addition of duckweed prevented weight loss. Further, in the presence of a high dose of Cu and Zn, DNA damage decreased, antioxidant gene expressions and enzymatic activities increased. In conclusion, results suggest that duckweed and Nile tilapia can be suitable candidates in metal remediation wastewater assessment programs.

Among post-translational modifications of proteins, ADP-ribosylation has been studied for over fifty years, assigning to this PTM a large set of functions, including DNA repair, transcription and cell signaling. This review presents an update on the function of a large set of enzyme writers, the readers that are recruited by the modified targets, and the erasers that reverse the modification to the original amino acid residue, removing the covalent bonds formed. In particular, the review provides details on the involvement of the enzymes performing MAR/PAR cycling in cancers. Of note, there is potential for application of the inhibitors developed for cancer also in the therapy of non-oncological diseases such as the protection against oxidative stress, suppression of inflammatory responses, and the treatment of neurodegenerative diseases. This field of studies is not concluded, since novel enzymes are being discovered at a rapid pace.