Browsing by Author "Dharmendra Kumar Chaudhary"

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Analysis of the multicopper oxidase gene regulatory network of Aeromonas hydrophila
(Kluwer Academic Publishers, 2012) Vijai Singh; Indra Mani; Dharmendra Kumar Chaudhary
Multicopper oxidase (MCO) is an enzyme which involves in reducing the oxygen in a four electron reduction to water with concomitant one electron oxidation of reducing the substrate. We have generated the 3-D structure of MCO by homology modeling and validated on the basis of free energy while 90. 4 % amino acid residues present in allowed regions of Ramachandran plot. The screening of potential hazardous aromatic compounds for MCO was performed using molecular docking. We obtained Sulfonaphthal, Thymolphthalein, Bromocresol green and Phloretin derivatives of phenol and aromatic hydrocarbon were efficient substrates for MCO. The phylogeny of MCO reveals that other bacteria restrain the homologous gene of MCO may play an important role in biodegradation of aromatic compounds. We have demonstrated the gene regulatory network of MCO with other cellular proteins which play a key role in gene regulation. These findings provide a new insight for oxidization of phenolic and aromatic compounds using biodegradation process for controlling environmental pollution. © 2012 Springer Science+Business Media B.V.
ATP4A gene regulatory network for fine-tuning of proton pump and ion channels
(2013) Vijai Singh; Indra Mani; Dharmendra Kumar Chaudhary
The ATP4A encodes α subunit of H+, K+-ATPase that contains catalytic sites of the enzyme forming pores through cell membrane which allows the ion transport. H+, K+-ATPase is a membrane bound P-type ATPase enzyme which is found on the surface of parietal cells and uses the energy derived from each cycle of ATP hydrolysis that can help in exchanging ions (H+, K+ and Cl-) across the cell membrane secreting acid into the gastric lumen. The 3-D model of α-subunit of H+, K+-ATPase was generated by homology modeling. It was evaluated and validated on the basis of free energies and amino acid residues. The inhibitor binding amino acid active pockets were identified in the 3-D model by molecular docking. The two drugs Omeprazole and Rabeprazole were found more potent interactions with generated model of α-subunit of H+, K+-ATPase on the basis of their affinity between drug-protein interactions. We have generated ATP4A gene regulatory networks for interactions with other proteins which involved in regulation that can help in fine-tuning of proton pump and ion channels. These findings provide a new dimension for discovery and development of proton pump inhibitors and gene regulation of the ATPase. It can be helpful in better understanding of human physiology and also using synthetic biology strategy for reprogramming of parietal cells for control of gastric ulcers. © 2013 Springer Science+Business Media Dordrecht.
Codon optimization of the major antigen encoding genes of diverse strains of influenza a virus
(International Association of Scientists in the International Association of Scientists in the, 2011) Indra Mani; Vijai Singh; Dharmendra Kumar Chaudhary; Pallavi Somvanshi; M.P.S. Negi
A large number of influenza A virus outbreaks and mortality occurred in the world recently, an urgent attention to develop effective and sufficient quantity of vaccines are needed. Vaccines are generally protein with immunogenic properties and are not expressed in sufficient quantity because of the codon bias, so it is necessary to optimize its codon in the expression host. Codon optimization was used to improve the protein expression in living organisms by increasing the translational efficiency of gene of interest. Two surface antigenic glycoproteins, hemagglutinin (HA) and neuraminidase (NA) are present in influenza A viruses. We have used HA and NA genes from 19 strains of influenza A viruses for codon optimization in E. coli. Both genes of the influenza virus show that the codon adaptation index (CAI) and GC content of the genes in optimized DNA were enhanced significantly (p <0.01) as compared to wild type. CAI and GC of HA in optimized DNA was enhanced by 3.2 (68.5%) and 1.2 (16.2%) fold respectively, while in NA it was increased by 3.3 (69.7%) and 1.2 (15.8%) fold respectively. Our finding demonstrates that the optimized genes could be useful for better expression in host without any truncated proteins and also helpful for protein folding and function. This work provides new insight in the synthetic biology research. © 2011 International Association of Scientists in the Interdisciplinary Areas and Springer-Verlag Berlin Heidelberg.
Development of diagnostic and vaccine markers through cloning, expression, and regulation of putative virulence-protein-encoding genes of Aeromonas hydrophila
(2013) Vijai Singh; Dharmendra Kumar Chaudhary; Indra Mani; Rohan Jain; B.N. Mishra
Aeromonas hydrophila is an opportunistic bacterial pathogen that is associated with a number of diseases in fish, amphibians, reptiles, and humans. In fish it causes several disease symptoms including tail and skin rot, and haemorrhagic septicemia; in human it causes soft-tissue wound infection and diarrhoea. The pathogenesis of A. hydrophila is multifactorial, but the mechanism is unknown so far. It is considered to be mediated by expression and secretion of extracellular proteins such as aerolysin, lipase, chitinase, amylase, gelatinase, hemolysins, and enterotoxins. A number of the putative virulence-protein-encoding genes that are present in the genome of A. hydrophila have been targeted by PCR for molecular diagnosis. These significant genes are also targeted for over-production of proteins by cloning and expression methods. In this review, we emphasize recent progress in the cloning, expression, and regulation of putative virulence-protein-encoding genes of A. hydrophila for a better understanding of the pathogenesis and also help to provide effective strategies for control of diseases. © 2013 The Microbiological Society of Korea and Springer-Verlag Berlin Heidelberg.
Gene cloning and homology modeling of the 3-oxoacyl-ACP synthase from Aeromonas hydrophila for drug discovery
(2011) Vijai Singh; Indra Mani; Dharmendra Kumar Chaudhary
Fatty acid biosynthesis in the Aeromonas hydrophila plays an important role in the formation of cell membrane and cell viability. The 3-oxoacyl- acyl carrier protein (ACP) synthase fadH and fadB from A. hydrophila have contributed in the initiation and elongation of fatty acid biosynthesis. In this study, we have designed new set of primers for amplification of the open reading frame of fadH and fadB. Cloning and sequencing of these amplified genes, analyzed that the G+C content of fadH and fadB were 63.3% and 68.79% respectively. The homology modeling was performed to generate the 3- D structure of fadH and fadB; and showed that more than 90% amino acid residues in allowed region of Ramachandranan plot. These findings provide a new avenue for better understanding, the role of genes encoding proteins involved in the fatty acid biosynthesis and thereby using as a potential and novel target for structure based drug designing to control the infections of A. hydrophila. © 2011 Bentham Science Publishers Ltd.
Gene network analysis of Aeromonas hydrophila for novel drug target discovery
(2012) Vijai Singh; Dharmendra Kumar Chaudhary; Indra Mani
Increasing the multi-drug resistance Aeromonas hydrophila creates a health problem regularly thus, an urgent needs to develop and screen potent antibiotics for controlling of the infections. There are many studies have focused on interactions between specific drugs, little is known about the system properties of a full drug interaction in gene network. Thus, an attractive approach for developing novel antibiotics against DNA gyrase, an enzyme essential for DNA replication, transcription, repair and recombination mechanisms which is important for bacterial growth and cell division. Homology modeling method was used to generate the 3-D structure of B subunit of DNA gyrase (gyrB) using known crystal structure. The active amino acids in 3-D structure of gyrB were targeted for structure based virtual screening of potent drugs by molecular docking. Number of drugs and analogs were selected and used for docking against gryB. The drugs Cinodine I, Cyclothialidine and Novobiocin were found to be more binding affinity with gyrB-drug interaction. The homology of gyrB protein sequence of A. hydrophila resembles with other species of Aeromonas closely showed relationship in phylogenetic tree. We have also demonstrated the gene network interactions of gyrB with other cellular proteins which are playing the key role in gene regulation. These findings provide new insight to understand the 3-D structure of gyrB which can be used in structure-based drug discovery; and development of novel, potent and specific drug against B subunit of DNA gyrase. © 2012 Springer Science+Business Media B.V.
Genotyping of Aeromonas hydrophila by Box elements
(2010) Vijai Singh; Dharmendra Kumar Chaudhary; Indra Mani; Pallavi Somvanshi; Gaurav Rathore; Neeraj Sood
PCR-based DNA fingerprinting techniques were evaluated to genotype eight diseased, particularly normal and environmental isolates of Aeromonas hydrophila. PCR-based fingerprinting method has an advantage of having repetitive sequence also called Box elements that are interspersed throughout the genome in diverse bacterial species. The BOX-PCR fingerprinting technique was evaluated for the discrimination of different isolates of A. hydrophila. All the studied isolates have shown major banding patterns ranged from 500-3000 bp. These finding could be advantageous to investigate the strain level specific fingerprints of A. hydrophila as potential genotypic markers. © 2010 Pleiades Publishing, Ltd.
Modification of histone by glyoxal: Recognition of glycated histone containing advanced glycation adducts by serum antibodies of type 1 diabetes patients
(Oxford University Press, 2018) Nadeem Ahmad Ansari; Dharmendra Kumar Chaudhary; Debabrata Dash
Dicarbonyl compounds react more rapidly, than glucose, with arginine and lysine in proteins to form advanced glycation end products (AGEs) and further produce free radicals which cause DNA damage. AGEs are reliable diagnostic biomarkers for most of the age-related diseases. In the present study histone was modified with glyoxal and it was characterized by various spectral techniques. Binding characteristics of the modified histone towards serum antibodies from type 1 diabetes patients was evaluated by solid phase enzyme immunoassay and the results were compared with normal human subjects. Fluorescence and Fourier transformed infrared analysis of the nuclear protein clearly indicated changes in their respective intensities upon modification with glyoxal. Liquid chromatography together with mass spectrometry showed new peaks and m/z values related to AGE adducts of dihydroimidazolidines/hydroimidazolones. This glyoxal modified protein was recognized by serum antibodies of the diabetes patients while it showed negligible binding with that of normal human subjects. Glyoxal modification of histone causes structural turbulence and formation of advanced glycation adducts in histone. These adducts might be the main antigenic epitope of the modified histone, leading to its recognition by circulating type 1 diabetes antibodies. © The Author(s) 2018. Published by Oxford University Press. All rights reserved.
Molecular detection and cloning of thermostable hemolysin gene from Aeromonas hydrophila
(2011) Vijai Singh; Indra Mani; Dharmendra Kumar Chaudhary; Pallavi Somvanshi
Aeromonas hydrophila is a major bacterial pathogen associated with hemorrhagic septicemia in aquatic and terrestrial animals including humans. There is an urgent need to develop molecular and immunological assays for rapid, specific and sensitive diagnosis. A new set of primers has been designed for detection of thermostable hemolysin (TH) gene (645 bp) from A. hydrophila, and sensitivity limit for detection of TH gene was 5 pg. The TH gene was cloned, sequenced and analyzed. The G+C content was 68.06%; and phylogeny was constructed using TH protein sequences which had significant homology with those for thermostable and other hemolysins present in several bacterial pathogens. In addition, we have predicted the four and eight T-cell epitopes for MHC class I and II alleles, respectively. These results provide new insight for TH protein containing antigenic epitopes that can be used in immunoassays and also designing of thermostable vaccines. © 2011 Pleiades Publishing, Ltd.