Subtractive genomics study for the identification of therapeutic targets against Cronobacter sakazakii: A threat to infants.

Cronobacter sakazakii is an opportunistic pathogen that has been associated with severe infection in neonates such as necrotizing enterocolitis (NEC), neonatal meningitis, and bacteremia. This pathogen can survive in a relatively dry environment, especially in powdered infant formula (PIF). Unfortunately, conventional drugs that were once effective against C. sakazakii are gradually losing their efficacy due to rising antibiotic resistance. In this study, a subtractive genomic approach was followed in order to identify potential therapeutic targets in the pathogen. The whole proteome of the pathogen was filtered through a step-by-step process, which involved removing paralogous proteins, human homologs, sequences that are less essential for survival, proteins with shared metabolic pathways, and proteins that are located in cells other than the cytoplasmic membrane. As a result, nine novel drug targets were identified. Further, the analysis also unveiled that the FDA-approved drug Terbinafine can be repurposed against the Glutathione/l-cysteine transport system ATP-binding/permease protein CydC of C. sakazakii. Moreover, molecular docking and dynamics studies of Terbinafine and CydC suggested that this drug can be used to treat C. sakazakii infection in neonates. However, for clinical purposes further in vitro and in vivo studies are necessary.

Investigation of pathogenic germline variants in gastric cancer and development of "GasCanBase" database.

BACKGROUND: Gastric cancer, which is also known as stomach cancer, can be influenced by both germline and somatic mutations. Non-synonymous Single Nucleotide Polymorphisms (nsSNPs) in germline have long been reported to play a pivotal role in cancer progression. AIM: The aim of this study is to examine the nsSNP in GC-associated genes. The study also aims to develop a database with extensive information regarding the nsSNPs in the GC-associated genes and their impacts. METHODS AND RESULTS: A total of 34,588 nsSNPs from 1,493,460 SNPs of the 40 genes were extracted from the available SNP database. Drug binding and energy minimization were examined by molecular docking and YASARA. To validate the existence of the germline CDH1 gene mutation (rs34466743) in the isolated blood DNA of gastric cancer (GC) patients, polymerase chain reaction (PCR) and DNA sequencing were performed. According to the results of the gene network analysis, 17 genes may interact with other types of cancer. A total of 11,363 nsSNPs were detected within the 40 GC genes. Among these, 474 nsSNPs were predicted to be damaging and 40 to be the most damaging. The SNPs in domain regions were thought to be strong candidates that alter protein functions. Our findings proposed that most of the selected nsSNPs were within the domains or motif regions. Free Energy Deviation calculation of protein structure pointed toward noteworthy changes in the structure of each protein that can demolish its natural function. Subsequently, drug binding confirmed the structural variation and the ineffectiveness of the drug against the mutant model in individuals with these germline variants. Furthermore, in vitro analysis of the rs34466743 germline variant from the CDH1 gene confirmed the strength and robustness of the pipeline that could expand the somatic alteration for causing cancer. In addition, a comprehensive gastric cancer polymorphism database named "GasCanBase" was developed to make data available to researchers. CONCLUSION: The findings of this study and the "GasCanBase" database may greatly contribute to our understanding of molecular epidemiology and the development of precise therapeutics for gastric cancer. GasCanBase is available at: https://www.gascanbase.com/.

High risk genetic variants of human insulin receptor substrate 1(IRS1) infer structural instability and functional interference.

Insulin receptor substrate 1(IRS1) is a signaling adapter protein encoded by the IRS1 gene. This protein delivers signals from insulin and insulin-like growth factor-1(IGF-1) receptors to the phosphatidylinositol 3-kinases (P13K)/protein kinase B (Akt) and Extracellular signal-regulated kinases (Erk) - Mitogen-activated protein (MAP) kinase pathways, which regulate particular cellular processes. Mutations in this gene have been linked to type 2 diabetes mellitus, a heightened risk of insulin resistance, and an increased likelihood of developing a number of different malignancies. The structure and function of IRS1 could be severely compromised as a result of single nucleotide polymorphism (SNP) type genetic variants. In this study, we focused on identification of the most harmful non-synonymous SNPs (nsSNPs) of the IRS1 gene as well as prediction of their structural and functional consequences. Six different algorithms made the initial prediction that 59 of the 1142 IRS1 nsSNPs would have a negative impact on the protein structure. In-depth evaluations detected 26 nsSNPs located inside the functional domains of IRS1. Following that, 16 nsSNPs were identified as more harmful based on conservation profile, hydrophobic interaction, surface accessibility, homology modelling, and inter-atomic interactions. Following an in-depth analysis of protein stability, M249T (rs373826433), I223T (rs1939785175) and V204G (rs1574667052) were identified as three most deleterious SNPs and were subjected to molecular dynamics simulation for further insights. These findings will help us understand the implications for disease susceptibility, cancer progression, and the efficacy of therapeutic development against IRS1 gene mutants.Communicated by Ramaswamy H. Sarma.

High-Risk Polymorphisms Associated with the Molecular Function of Human HMGCR Gene Infer the Inhibition of Cholesterol Biosynthesis.

HMG-CoA reductase or HMGCR (3-hydroxy-3-methylglutaryl-CoA reductase) is a rate-limiting enzyme involved in cholesterol biosynthesis. HMGCR plays an important role in the possible occurrence of hypercholesterolemia leading to atherosclerosis and coronary heart disease. This enzyme is a major target for cholesterol-lowering drugs such as "statin" which blocks the synthesis of mevalonate, a precursor for cholesterol biosynthesis. This study is aimed at characterizing deleterious mutations and classifying functional single nucleotide polymorphisms (SNPs) of the HMGCR gene through analysis of functional and structural evaluation, domain association, solvent accessibility, and energy minimization studies. The functional and characterization tools such as SIFT, PolyPhen, SNPs and GO, Panther, I-Mutant, and Pfam along with programming were employed to explore all the available SNPs in the HMGCR gene in the database. Among 6815 SNP entries from different databases, approximately 388 SNPs were found to be missense. Analysis showed that seven missense SNPs are more likely to have deleterious effects. A tertiary model of the mutant protein was constructed to determine the functional and structural effects of the HMGCR mutation. In addition, the location of the mutations suggests that they may have deleterious effects because most of the mutations are residing in the functional domain of the protein. The findings from the analysis predicted that rs147043821 and rs193026499 missense SNPs could cause significant structural and functional instability in the mutated proteins of the HMGCR gene. The findings of the current study will likely be useful in future efforts to uncover the mechanism and cause of hypercholesterolemia. In addition, the identified SNPs of HMGCR gene could set up a strong foundation for further therapeutic discovery.

Effect of gamma radiation on microbial load, physico-chemical and sensory characteristics of common spices for storage.

The effect of gamma radiation on the decontamination of microbial population, physico-chemical, radiation sensitivity and sensory characteristics of common spices for storage were evaluated. Spices were irradiated with gamma doses of 0 (as control), 2, 4, 6, 8 and 10 kGy, packed in the glass vials and stored at room temperature (22 ± 2°C) in the laboratory. In this research, Bacillus, Salmonella and Listeria species were identified in un-irradiated spice samples. Results also indicated that gamma radiation reduced the total microbial population compared to control and optimum gamma radiation doses (6 kGy for red chili and turmeric; 4 kGy for cumin, coriander, garlic and black pepper; 2 kGy for ginger powder samples) were identified for decontamination of the organisms in the studied spices. It was concluded that no significant differences before and after gamma radiation were observed in physico-chemical, nutritional and sensory properties but significantly changed in microbial load in spices samples.

Insight of druggable cannabinoids against estrogen receptor ß in breast cancer.

Breast cancer (BC) is the second most prevalent cancer worldwide. Estrogen receptor beta (ERß) is an essential protein of breast cells to suppress estrogen-induced uncontrolled proliferation. Thus, small molecules that can modulate and enhance ERß expression would be an effective agent to suppress BC development. Studies showed that cannabinoid (CB), specifically delta-9-tetrahydrocannabinol (Del9THC), can increase the expression of ERß and inhibits BC cell proliferation. In this study, less psychoactive and structurally similar analogs of Del9THC were chosen as drug candidates and ERß was targeted as a therapeutic receptor. Delta-8-tetrahydrocannabinol (Del8THC) and delta-4-isotetrahydrocannabinol (Del4isoTHC) were the drug candidates selected on the basis of literature reports, absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties, medicinal chemistry profile, and physicochemical features. Molecular docking simulations were carried out to determine ligand receptor interactions and binding affinity based on free binding energy. To get a better drug, the structural modification was done on Del8THC and generated a new CB analog called Cannabinoid A. Finally, molecular interaction analysis revealed that two CBs and one of their analog interact with the active site residues of ERß. Therefore, this study revealed a new way to discover novel drug(s) for BC patients.Communicated by Ramaswamy H. Sarma.

Molecular evolution and genomics of hepatitis B virus subgenotype C2 strain predominant in the chronic patients in Bangladesh.

Evolution of hepatitis B virus (HBV) is a mystery and caused mainly by genomic mutations as well as recombination. Viral evolution may be responsible for increasing disease severity and render resistance to the existing treatment processes. HBV/C2 strain is associated with chronicity, which may progress to the liver cirrhosis and hepatocellular carcinoma. Furthermore, HBV/C2 strain is highly prevalent in the chronic hepatitis B patients in Bangladesh. Hence, the molecular evolution of that strain and its disease pattern need to be uncovered. Herein, the purpose of this study is to determine the potential mutations of HBV complete genome sequences isolated in Bangladesh and the molecular evolution of HBV/C2 strain. Mutation analysis of the total 57 complete genome sequences of HBV in Bangladesh revealed that 42.11%, 12.28%, 7.02% and 3.51% of the strains were vaccine resistant, HBsAg detection escape, HBV immunoglobulin escape, multi-drug resistant respectively. Furthermore, of the vaccine resistant strains, 16.67% were observed resistant to both vaccine, HBsAg detection and immunoglobulin escape. Bayesian skyline analysis with 462 HBV/C2 strains from 2000 to 2017 revealed the evolution of the strain was in nineteenth century with two rapid sharp increases in the genetic diversity at the end of the twentieth century and then a sudden decrease in the early twenty-first century as observed in C and X gene analysis. This study may help researchers and clinicians to get a better knowledge about the emergence and evolution of HBV/C2 strain that may help to find a proper treatment strategy against hepatitis B.

High-throughput three-dimensional gel electrophoresis for versatile utilities: a stacked slice-gel system for separation and reactions (4SR).

A novel high-throughput system, called the stacked slice-gel system for separation and reactions (4SR), was developed for the analysis of DNA/RNA and protein/peptide. The system provides a novel three-dimensional gel electrophoresis approach that exploits the property of stacked slice gels. It allows multiple samples simultaneously to react as well as to be separated, offering a two-dimensional (mxn) sample loading system. For this purpose, high-throughput multi-micro vessels (MMVs) containing variable numbers of wells (100 wells in this paper) have been used, which are made of 25 mm square-size polyacrylamide gels. Furthermore, after electrophoretic separation, a slice gel containing a desired sample can be easily removed and proceeded to the next step. Different biological reactions as well as successive separation of products were effectively carried out dealing with DNA/RNA and protein/peptide. It shows that this system has a diversity of potentials to be developed.

Efficient SNP analysis enabled by joint application of the muTGGE and heteroduplex methods.

Gene science-based diagnoses have become an increasingly realistic option as the state of knowledge has improved regarding the genetic basis of disease. To facilitate the creation of this potential diagnostic tool, researchers have made large-scale detection of point mutations a key issue. Here, we propose an inexpensive and convenient method with a high performance level for this purpose: micro temperature gradient gel electrophoresis (muTGGE)-empowered heteroduplex analysis (muTG-HD). First, muTGGE was shown to separate double-stranded DNA containing single nucleotide polymorphism (SNP) with sufficiently high resolution when used in the mode of perpendicular TGGE. Using human c-Ki-ras and rat p53 DNA, point mutations could be unequivocally detected by muTG-HD when parallel TGGE was employed. The mutation type (such as G/C to A/T), the position of the point mutation (centre or not) and the DNA size (around 100 or 200 bp) were examined and found to be detectable. Thus, muTG-HD could detect point mutations efficiently at a much lower cost by having multiple lanes per gel.