Publications by authors named "Nar Singh Chauhan"

20 Publications

  • Page 1 of 1

Mapping of the benzoate metabolism by human gut microbiome indicates food-derived metagenome evolution.

Sci Rep 2021 Mar 10;11(1):5561. Epub 2021 Mar 10.

Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana, 124001, India.

Sodium benzoate is one of the widely used food preservatives and its metabolism in the human body has been studied only with the host perspective. Despite the human gut microbiome being considered as a virtual human organ, its role in benzoate metabolism is yet to be elucidated. The current study uses a multi-omic approach to rationalize the role of human gut microbes in benzoate metabolism. Microbial diversity analysis with multiple features synchronously indicates the dominance of Bacteroidetes followed by Firmicutes, Actinobacteria, and Proteobacteria. Metagenomic exploration highlights the presence of benzoate catabolic protein features. These features were mapped on to the aerobic and anaerobic pathways of benzoate catabolism. Benzoate catabolism assays identified statistically significant metabolites (P < 0.05) associated with the protocatechuate branch of the beta-ketoadipate pathway of the benzoate metabolism. Analysis of the 201 human gut metagenomic datasets across diverse populations indicates the omnipresence of these features. Enrichment of the benzoate catabolic protein features in human gut microbes rationalizes their role in benzoate catabolism, as well as indicates food-derived microbiome evolution.
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http://dx.doi.org/10.1038/s41598-021-84964-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7946887PMC
March 2021

Comparative analysis of the alveolar microbiome in COPD, ECOPD, Sarcoidosis, and ILD patients to identify respiratory illnesses specific microbial signatures.

Sci Rep 2021 Feb 17;11(1):3963. Epub 2021 Feb 17.

Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana, 124001, India.

Studying respiratory illness-specific microbial signatures and their interaction with other micro-residents could provide a better understanding of lung microbial ecology. Each respiratory illness has a specific disease etiology, however, so far no study has revealed disease-specific microbial markers. The present study was designed to determine disease-specific microbial features and their interactions with other residents in chronic obstructive pulmonary diseases (stable and exacerbated), sarcoidosis, and interstitial lung diseases. Broncho-alveolar lavage samples (n = 43) were analyzed by SSU rRNA gene sequencing to study the alveolar microbiome in these diseases. A predominance of Proteobacteria followed by Firmicutes, Bacteroidetes, Actinobacteria, and Fusobacteria was observed in all the disease subsets. Shannon diversity was significantly higher in stable COPD when compared to exacerbated chronic obstructive pulmonary disease (ECOPD) (p = 0.0061), and ILD patient samples (p = 0.037). The lung microbiome of the patients with stable COPD was more diverse in comparison to ECOPD and ILD patients (p < 0.001). Lefse analysis identified 40 disease-differentiating microbial features (LDA score (log10) > 4). Species network analysis indicated a significant correlation (p < 0.05) of diseases specific microbial signature with other lung microbiome members. The current study strengthens the proposed hypothesis that each respiratory illness has unique microbial signatures. These microbial signatures could be used as diagnostic markers to differentiate among various respiratory illnesses.
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http://dx.doi.org/10.1038/s41598-021-83524-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7889618PMC
February 2021

Catabolic Machinery of the Human Gut Microbes Bestow Resilience Against Vanillin Antimicrobial Nature.

Front Microbiol 2020 16;11:588545. Epub 2020 Oct 16.

Department of Biochemistry, Maharshi Dayanand University, Rohtak, India.

Vanillin is a phenolic food additive commonly used for flavor, antimicrobial, and antioxidant properties. Though it is one of the widely used food additives, strategies of the human gut microbes to evade its antimicrobial activity await extensive elucidation. The current study explores the human gut microbiome with a multi-omics approach to elucidate its composition and metabolic machinery to counter vanillin bioactivity. A combination of SSU rRNA gene diversity, metagenomic RNA features diversity, phylogenetic affiliation of metagenome encoded proteins, uniformly ( = 0.99) indicates the abundance of Bacteroidetes followed by Firmicutes and Proteobacteria. Manual curation of metagenomic dataset identified gene clusters specific for the vanillin metabolism (, and ) and intermediary metabolic pathways ( and operon). Metagenomic dataset comparison identified the omnipresence of vanillin catabolic features across diverse populations. The metabolomic analysis brings forth the functionality of the vanillin catabolic pathway through the Protocatechuate branch of the beta-ketoadipate pathway. These results highlight the human gut microbial features and metabolic bioprocess involved in vanillin catabolism to overcome its antimicrobial activity. The current study advances our understanding of the human gut microbiome adaption toward changing dietary habits.
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http://dx.doi.org/10.3389/fmicb.2020.588545DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7605359PMC
October 2020

Hypoxia Inducible Factor-1α: The Curator of Gut Homeostasis.

Front Cell Infect Microbiol 2020 15;10:227. Epub 2020 May 15.

Department of Biochemistry, Maharshi Dayanand University, Rohtak, India.

The human gut microbiome is a stratified and resilient ecosystem co-inhabited by a diverse and dynamic pool of microorganisms. Microbial selection, establishment, and colonization are modulated through a complex molecular network of host-microbial interactions. These molecular bioprocesses ensure the taxonomic composition of the mature human gut microbiome. The human gut microbiome plays a vital role in host health; otherwise, any microbial dysbiosis could predispose to the onset of physiological and metabolic disorder/s. Focussed research are being carried out to identify key molecular agents defining gut homeostasis. These molecules hold the potential to develop effective therapeutic solutions for microbial dysbiosis-associated human disorders. Of these, Hypoxia-inducible factor-1α (HIF-1α) is a central player in host-microbial crosstalk to maintain gut homeostasis. Human gut microbial metabolites regulate its cellular stability, which in turn regulates various cellular processes required for the stable gut microbiome. In the present review, an effort has been made to summarize the key role of HIF-1α to maintain gut homeostasis.

Highlights: - Explain the molecular process of host microbial molecular interactions.- Establish the explicit role of HIF-1α in intestinal epithelial integrity and gut health.- Regulation of HIF-1α by human gut commensals and vice a versa.- Regulation of the host immune response for survival and colonization of human gut commensal.
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http://dx.doi.org/10.3389/fcimb.2020.00227DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7242652PMC
May 2020

Curcumin Encapsulated PEGylated Nanoliposomes: A Potential Anti-Infective Therapeutic Agent.

Indian J Microbiol 2019 Sep 7;59(3):336-343. Epub 2019 Jun 7.

1Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana India.

Exploration of novel bioactive molecules or potentiation of the existing bioactive molecules is necessary to reduce the burden of the infectious diseases for the better human health. Curcumin is a promising molecule with huge therapeutic potential. Despite high bioactivity, its therapeutic suitability is shadowed by poor bioavailability, limited aqueous solubility, and short shelf life. Nanotechnology has generated new avenues to overcome these challenges. In the current study polymer assisted nanoliposomes, PEGylated Curcumin nanoliposomes with good loading efficiency were prepared. These particles have shown 1000 fold enhanced curcumin hydrophilicity and tenfold higher stability. In vitro release kinetic indicates two fold higher curcumin release in the simulated gastric and intestinal environment. Various bioactivity assays have confirmed enhanced bioactivity of nanocurcmin in comparison of the native curcumin. PEGylated Curcumin nanoliposomes could be employed for treating various diseases.
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http://dx.doi.org/10.1007/s12088-019-00811-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6646638PMC
September 2019

S9A Serine Protease Engender Antigenic Gluten Catabolic Competence to the Human Gut Microbe.

Indian J Microbiol 2018 Sep 27;58(3):294-300. Epub 2018 Apr 27.

1Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana 124001 India.

The human gut microbiome has a significant role in host physiology; however its role in gluten catabolism is debatable. Present study explores the role of human gut microbes in gluten catabolism and a native human gut microbe sp. HM71 was identified. SSU rDNA analysis has described human gut microbiome structure and also confirmed the permanent residentship of sp. HM71. Catabolic potential of sp. HM71 to cleave antigenic gluten peptides indicates presence of candidate gene encoding biocatalytic machinery. Genome analysis has identified the presence of gene encoding S9A serine protease family-prolyl endopeptidase, with Ser591, Asp664 and His685 signature residues. sp. HM71 prolyl endopeptidase activity was found optimal at pH 7.0 and 37 °C with a of 35.53 μmol and specifically cleaves at proline residue. Current study describes the gluten catabolism potential of sp. HM71 depicting possible role of human gut microbes in gluten catabolism to confer resistance mechanisms for the onset of celiac diseases in populations with gluten diet.
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http://dx.doi.org/10.1007/s12088-018-0732-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6023808PMC
September 2018

TiO and its composites as promising biomaterials: a review.

Biometals 2018 04 1;31(2):147-159. Epub 2018 Feb 1.

Department of Chemistry, Maharshi Dayanand University, Rohtak, India.

TiO is a well-known material and has remarkable physical, chemical and biocompatible properties which have made it a suitable material in the biological world. The development of new TiO-based materials is strongly required to achieve desired properties and applications. A large number of TiO composites have been synthesized and applied in various fields. The present review reports the utility of TiO and its composites in biosensing, in Photodynamic Therapy, as an antimicrobial agent and as a nanodrug carrier. The aim of this review is to discuss the biological application of the TiO based materials and some recent advancement in TiO to enhance its application in the biological world.
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http://dx.doi.org/10.1007/s10534-018-0078-6DOI Listing
April 2018

Functional metagenomics identifies novel genes ABCTPP, TMSRP1 and TLSRP1 among human gut enterotypes.

Sci Rep 2018 01 23;8(1):1397. Epub 2018 Jan 23.

Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana, India.

Every niche in the biosphere is touched by the seemingly endless capacity of microbes to transform the world around them by adapting swiftly and flexibly to the environmental changes, likewise the gastrointestinal tract is no exception. The ability to cope with rapid changes in external osmolarity is an important aspect of gut microbes for their survival and colonization. Identification of these survival mechanisms is a pivotal step towards understanding genomic suitability of a symbiont for successful human gut colonization. Here we highlight our recent work applying functional metagenomics to study human gut microbiome to identify candidate genes responsible for the salt stress tolerance. A plasmid borne metagenomic library of Bacteroidetes enriched human fecal metagenomic DNA led to identification of unique salt osmotolerance clones SR6 and SR7. Subsequent gene analysis combined with functional studies revealed that TLSRP1 within pSR7 and TMSRP1 and ABCTPP of pSR6 are the active loci responsible for osmotolerance through an energy dependent mechanism. Our study elucidates the novel genetic machinery involved in bestowing osmotolerance in Prevotella and Bacteroidetes, the predominant microbial groups in a North Indian population. This study unravels an alternative method for imparting ionic stress tolerance, which may be prevalent in the human gut microbiome.
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http://dx.doi.org/10.1038/s41598-018-19862-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5780487PMC
January 2018

A review of metabolic potential of human gut microbiome in human nutrition.

Arch Microbiol 2018 Mar 29;200(2):203-217. Epub 2017 Nov 29.

Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana, 124001, India.

The human gut contains a plethora of microbes, providing a platform for metabolic interaction between the host and microbiota. Metabolites produced by the gut microbiota act as a link between gut microbiota and its host. These metabolites act as messengers having the capacity to alter the gut microbiota. Recent advances in the characterization of the gut microbiota and its symbiotic relationship with the host have provided a platform to decode metabolic interactions. The human gut microbiota, a crucial component for dietary metabolism, is shaped by the genetic, epigenetic and dietary factors. The metabolic potential of gut microbiota explains its significance in host health and diseases. The knowledge of interactions between microbiota and host metabolism, as well as modification of microbial ecology, is really beneficial to have effective therapeutic treatments for many diet-related diseases in near future. This review cumulates the information to map the role of human gut microbiota in dietary component metabolism, the role of gut microbes derived metabolites in human health and host-microbe metabolic interactions in health and diseases.
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http://dx.doi.org/10.1007/s00203-017-1459-xDOI Listing
March 2018

Identification of Arsenic Resistance Genes from Marine Sediment Metagenome.

Indian J Microbiol 2017 Sep 5;57(3):299-306. Epub 2017 Jul 5.

CSIR-Institute of Genomics and Integrative Biology, Council of Scientific and Industrial Research, Sukhdev Vihar, Mathura Road, Delhi, 110020 India.

A metagenomic library of sea sediment metagenome containing 245,000 recombinant clones representing ~ 2.45 Gb of sea sediment microbial DNA was constructed. Two unique arsenic resistance clones, A7 and A12, were identified by selection on sodium arsenite containing medium. Clone A7 showed a six-fold higher resistance to arsenate [As(V)], a three-fold higher resistance to arsenite [As(III)] and significantly increased resistance to antimony [Sb(III)], while clone A12 showed increased resistance only to sodium arsenite and not to the other two metalloids. The clones harbored inserts of 8.848 Kb and 6.771 Kb, respectively. Both the clones possess A + T rich nucleotide sequence with similarity to sequences from marine psychrophilic bacteria. Sequence and transposon-mutagenesis based analysis revealed the presence of a putative arsenate reductase (ArsC), a putative arsenite efflux pump (ArsB/ACR) and a putative NADPH-dependent FMN reductase (ArsH) in both the clones and also a putative transcriptional regulatory protein (ArsR) in pA7. The increased resistance of clone A7 to As(V), As(III) and Sb(III) indicates functional expression of ArsC and ArsB proteins from pA7. The absence of increased As(V) resistance in clone A12 may be due to the expression of a possible inactive ArsC, as conserved Arg60 residue in this protein was replaced by Glu60, while the absence of Sb(III) resistance may be due to the presence of an ACR3p-type arsenite pump, which is known to lack antimony transport ability.
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http://dx.doi.org/10.1007/s12088-017-0658-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5574776PMC
September 2017

Comparative Genomics of Host-Symbiont and Free-Living Oceanobacillus Species.

Genome Biol Evol 2017 05;9(5):1175-1182

Department of Biochemistry, Maharshi Dayanand University, Rohtak, Haryana, India.

Survival in a given environment requires specific functions, so genomic variation is anticipated within in individual taxonomic groups that exhibit a large diversity in lifestyles. In this study, we sequence and assemble the genome of Oceanobacillus faecalis strain HM6, a resident of the human gut. Using the genus Oceanobacillus and the HM6 draft genome sequence, we explore the functional requirements for survival in a symbiotic arrangement within the human gut, in contrast to free living in the environment. Comparative genomics of seven available Oceanobacillus complete genomes highlight a genomically heterogeneous group. Our analysis did not find strict phylogenetic separation between free-living and host-symbiont Oceanobacillus members. By comparing functional gene content between host-associated and free-living species, we identified candidate genes that are potentially involved in symbiotic lifestyles, including phosphotransferase genes, transporters and two component response regulators. This study summarizes genomic and phylogenetic differences in the Oceanobacillus genus. Additionally, we highlight functions that may be key for survival in the human gut community.
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http://dx.doi.org/10.1093/gbe/evx076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5425236PMC
May 2017

Physiopathology and Management of Gluten-Induced Celiac Disease.

J Food Sci 2017 Feb 31;82(2):270-277. Epub 2017 Jan 31.

Dept. of Biochemistry, M.D. Univ., Rohtak, 124001, Haryana, India.

Proline- and glutamine-rich gluten proteins are one of the major constituents of cereal dietary proteins, which are largely resistant to complete cleavage by the human gastrointestinal (GI) digestive enzymes. Partial digestion of gluten generates approximately 35 amino acids (aa) immunomodulatory peptides which activate T-cell-mediated immune system, followed by immunological inflammation of mucosa leading to the onset of celiac disease (CD). CD is an autoimmune disease associated with HLA-DQ2/DQ8 polymorphism and dysbiosis of gut microbiota. CD is either diagnosed using duodenal mucosal biopsis or serological testing for transglutaminase 2 (TG2) specific antibodies (IgA and IgG). Current therapy for CD management is gluten-free diet, while other therapies like glutenase, probiotics, immunomodulation, jamming of HLA-DQ2, inhibition of TG2, and gluten tolerance aided by gluten tolerizing vaccines are being developed.
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http://dx.doi.org/10.1111/1750-3841.13612DOI Listing
February 2017

An Improved Methodology to Overcome Key Issues in Human Fecal Metagenomic DNA Extraction.

Genomics Proteomics Bioinformatics 2016 Dec 23;14(6):371-378. Epub 2016 Nov 23.

Department of Biochemistry, Maharshi Dayanand University, Rohtak 124001, India. Electronic address:

Microbes are ubiquitously distributed in nature, and recent culture-independent studies have highlighted the significance of gut microbiota in human health and disease. Fecal DNA is the primary source for the majority of human gut microbiome studies. However, further improvement is needed to obtain fecal metagenomic DNA with sufficient amount and good quality but low host genomic DNA contamination. In the current study, we demonstrate a quick, robust, unbiased, and cost-effective method for the isolation of high molecular weight (>23kb) metagenomic DNA (260/280 ratio >1.8) with a good yield (55.8±3.8ng/mg of feces). We also confirm that there is very low human genomic DNA contamination (eubacterial: human genomic DNA marker genes=2:1) in the human feces. The newly-developed method robustly performs for fresh as well as stored fecal samples as demonstrated by 16S rRNA gene sequencing using 454 FLX+. Moreover, 16S rRNA gene analysis indicated that compared to other DNA extraction methods tested, the fecal metagenomic DNA isolated with current methodology retains species richness and does not show microbial diversity biases, which is further confirmed by qPCR with a known quantity of spike-in genomes. Overall, our data highlight a protocol with a balance between quality, amount, user-friendliness, and cost effectiveness for its suitability toward usage for culture-independent analysis of the human gut microbiome, which provides a robust solution to overcome key issues associated with fecal metagenomic DNA isolation in human gut microbiome studies.
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http://dx.doi.org/10.1016/j.gpb.2016.06.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5200916PMC
December 2016

Synthesis, characterization of penicillin G capped silver nanoconjugates to combat β-lactamase resistance in infectious microorganism.

J Biotechnol 2013 Feb 7;163(4):419-24. Epub 2013 Jan 7.

Department of Biochemistry, Maharshi Dayanand University, Rohtak 124001, Haryana, India.

In the present study, a novel strategy was adopted to synthesize, β lactamase resistant penicillin G molecules by using the unique properties of silver nanoparticles. Ascorbic acid-stabilized spherical monodispersed silver nanoparticles were prepared by a simple chemical reaction. The formation processes of the silver nanoparticles were investigated by UV-vis spectroscopy and Atomic Force Microscopy (AFM). Free amine groups were introduced on the surface of native silver nanoparticles by coating a uniform layer of polyaniline and this was confirmed by FTIR spectroscopy and Scanning Electron Microscopy. Functionalized silver nanoparticles were then grafted to the C3 carboxyl group of the β lactam ring of penicillin G in the presence of N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDAC) with a conjugation yield of 213 μg mg⁻¹. These novel silver penicillin G nanoconjugates showed a very good growth inhibition against both non-resistant Escherichia coli (gram negative) as well as toward β lactam resistant, E. coli (gram negative) and Staphylococcus aureus (gram positive).
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http://dx.doi.org/10.1016/j.jbiotec.2012.12.002DOI Listing
February 2013

Genome sequence of Rheinheimera sp. strain A13L, isolated from Pangong Lake, India.

J Bacteriol 2011 Oct 8;193(20):5873-4. Epub 2011 Jul 8.

Institute of Genomics and Integrative Biology, Council of Scientific and Industrial Research, Mall Road, Delhi 110007, India.

Rheinheimera sp. strain A13L, which has antimicrobial activity, was isolated from alkaline brackish water of the high-altitude Pangong Lake of Ladakh, India. Here we report the draft genome sequence of Rhienheimera sp. strain A13L (4,523,491 bp with a G+C content of 46.23%). The genome is predicted to contain genes for marinocine and colicin V production, which may be responsible for the antimicrobial activity of the strain.
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http://dx.doi.org/10.1128/JB.05636-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3187196PMC
October 2011

Cloning and characterization of an epoxide hydrolase from Cupriavidus metallidurans-CH34.

Protein Expr Purif 2011 Sep 15;79(1):49-59. Epub 2011 Apr 15.

Department of Biochemistry, Panjab University, Chandigarh, India.

A putative epoxide hydrolase-encoding gene was identified from the genome sequence of Cupriavidus metallidurans CH34. The gene was cloned and overexpressed in Escherichia coli with His(6)-tag at its N-terminus. The epoxide hydrolase (CMEH) was purified to near homogeneity and was found to be a homodimer, with subunit molecular weight of 36 kDa. The CMEH had broad substrate specificity as it could hydrolyze 13 epoxides, out of 15 substrates tested. CMEH had high specific activity with 1,2-epoxyoctane, 1,2-epoxyhexane, styrene oxide (SO) and was also found to be active with meso-epoxides. The enzyme had optimum pH and temperature of 7.5 and 37°C respectively, with racemic SO. Biotransformation of 80 mM SO with recombinant whole E. coli cells expressing CMEH led to 56% ee(P) of (R)-diol with 77.23% conversion in 30 min. The enzyme could hydrolyze (R)-SO, ∼2-fold faster than (S)-SO, though it accepted both (R)- and (S)-SO with similar affinity as K(m)(R) and K(m)(S) of CMEH were 2.05±0.42 and 2.11±0.16 mM, respectively. However, the k(cat)(R) and k(cat)(S) for the two enantiomers of SO were 4.80 and 3.34 s(-1), respectively. The wide substrate spectrum exhibited by CMEH combined with the fast conversion rate makes it a robust biocatalyst for industrial use. Regioselectivity studies with enantiopure (R)- and (S)-SO revealed that with slightly altered regioselectivity, CMEH has a high potential to synthesize an enantiopure (R)-PED, through an enantioconvergent hydrolytic process.
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http://dx.doi.org/10.1016/j.pep.2011.04.007DOI Listing
September 2011

Identification of two flavin monooxygenases from an effluent treatment plant sludge metagenomic library.

Bioresour Technol 2010 Nov 29;101(21):8481-4. Epub 2010 Jun 29.

Microbial Genomics and Biotechnology Group, Institute of Genomics and Integrative Biology, Council of Scientific and Industrial Research (CSIR), Mall Road, New Delhi 110007, India.

Oxygenases are useful for the production of many industrially important molecules. Screening of an effluent treatment plant (ETP) sludge metagenomic library identified two clones encoding proteins, B1 and B2, with similarity to putative flavin monooxygenases from Mesorhizobium loti and Sphingomonas wittichi, respectively. The deduced amino acid sequences show only 20% identity, but both have a paired Rossman fold and a flavin monooxygenase (FMO) motif. B1 and B2 appear to be members of the flavin-containing monooxygenase and the Baeyer-Villiger monooxygenases subfamilies, respectively. When expressed in Escherichia coli, the two clones produced activities that oxidized indole to a mixture of indigo and indirubin pigments. These results suggest that B1 and B2 have potential as a biocatalyst in indigo/indirubin production.
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http://dx.doi.org/10.1016/j.biortech.2010.06.025DOI Listing
November 2010

Chemical activation of egg shell membrane for covalent immobilization of enzymes and its evaluation as inert support in urinary oxalate determination.

Talanta 2009 Mar 14;77(5):1688-93. Epub 2008 Oct 14.

Biochemistry Research laboratory, Department of Biochemistry & Genetics, MD University, Rohtak 124001, Haryana, India.

Egg shell membrane is a novel, robust, microporous, cost effective, easily available organic support material. In recent studies egg shell membranes were utilized as organic support for enzyme immobilization. But low conjugation yield limits its application as good support for biotechnological industries. In present study egg shell membrane was chemically treated to introduce free functional groups for covalent linkage of proteins to increase its conjugation yield and stability of conjugate complex. Many enzymes were tested for immobilization on modified egg shell membrane like oxalate oxidase, glucose oxidase, peroxidase and lipase. A fifteen to sixteen fold increase in conjugation yield was observed when immobilization was performed after chemical treatment in comparison to immobilization on native membrane with slight change in specific activity of immobilized enzyme which ranges from 5% to 15%. Egg shell membrane bound enzymes showed slight changes in their kinetic properties after immobilization. Egg shell membrane bound oxalate oxidase shows detection limit of 1.5 microM when used for urinary oxalate determination. Egg shell membrane support shows no interference to enzyme activity and a good correlation of 0.99 was observed with the values estimated using commercially available Sigma kit. The immobilized oxalate oxidase, glucose oxidase, peroxidase and lipase were stable up to duration of 180 days and there is respective loss of 10%, 13%, 24%, and 33% of initial activity. Overall result strengthens our view of using chemically modified egg shell membrane as solid support for better immobilization of enzymes and can be used in various biotechnological applications.
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http://dx.doi.org/10.1016/j.talanta.2008.10.004DOI Listing
March 2009

Identification of genes conferring arsenic resistance to Escherichia coli from an effluent treatment plant sludge metagenomic library.

FEMS Microbiol Ecol 2009 Jan 4;67(1):130-9. Epub 2008 Nov 4.

Microbial Genomics and Biotechnology Group, Institute of Genomics and Integrative Biology, Council of Scientific and Industrial Research (CSIR), Delhi, India.

The majority of bacteria elude culture in the laboratory. A metagenomic approach provides culture-independent access to the gene pool of the whole bacterial community. A metagenomic library was constructed from an industrial effluent treatment plant sludge containing about 1.25 Gb of microbial community DNA. Two arsenic-resistant clones were selected from the metagenomic library. Clones MT3 and MT6 had eight- and 18-fold higher resistance to sodium arsenate in comparison with the parent strain, respectively. The clones also showed increased resistance to arsenite but not to antimony. Sequence analysis of the clones revealed genes encoding for putative arsenate reductases and arsenite efflux pumps. A novel arsenate resistance gene (arsN) encoding a protein with similarity to acetyltransferases was identified from clone MT6. ArsN homologues were found to be closely associated with arsenic resistance genes in many bacterial genomes. ArsN homologues were found fused to putative arsenate reductases in Methylibium petroleiphilum PM1 and Anaeromyxobacter dehalogenans 2CP-C and with a putative arsenite chaperone in Burkholderia vietnamiensis G4. ArsN alone resulted in an approximately sixfold higher resistance to sodium arsenate in wild-type Escherichia coli W3110.
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http://dx.doi.org/10.1111/j.1574-6941.2008.00613.xDOI Listing
January 2009

Activation of polyvinyl chloride sheet surface for covalent immobilization of oxalate oxidase and its evaluation as inert support in urinary oxalate determination.

Anal Biochem 2008 Mar 13;374(2):272-7. Epub 2007 Nov 13.

Biochemistry Research Laboratory, Department of Biochemistry and Genetics, MD University, Rohtak, 124001 Haryana, India.

Polyvinyl chloride (PVC) sheets are a promising material for enzyme immobilization owing to the PVC's properties such as being chemically inert, corrosion free, weather resistant, tough, lightweight, and maintenance free and having a high strength-to-weight ratio. In this study, this attractive material surface was chemically modified and exploited for covalent immobilization of oxalate oxidase using glutaraldehyde as a coupling agent. The enzyme was immobilized on activated PVC surface with a conjugation yield of 360 microg/cm(2). The scanning electron micrographs showed the microstructures on the PVC sheet surface revealing the successful immobilization of oxalate oxidase. A colorimetric method was adopted in evaluating enzymatic activity of immobilized and native oxalate oxidase. The immobilized enzyme retained 65% of specific activity of free enzyme. Slight changes were observed in the optimal pH, incubation temperature, and time for maximum activity of immobilized oxalate oxidase. PVC support showed no interference when immobilized oxalate oxidase was used for estimation of oxalic acid concentration in urine samples and showed a correlation of 0.998 with the values estimated with a commercially available Sigma kit. The overall results strengthen our view that PVC sheet can be used as a solid support for immobilization of enzymes and in the field of clinical diagnostics, environmental monitoring and remediation.
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http://dx.doi.org/10.1016/j.ab.2007.11.008DOI Listing
March 2008