Classification and functional analyses of putative virulence factors of Mycobacterium tuberculosis: A combined sequence and structure based study

The emergence of the drug-resistant mechanisms in Mycobacterium tuberculosis poses the biggest challenges to the current therapeutic measures, which necessitates the identification of new drug targets. The Hypothetical Proteins (HPs), a class of functionally uncharacterized proteins, may provide a new class of undiscovered therapeutic targets. The genome of M. tuberculosis contains 1000 HPs with their sequences were analyzed using a variety of bioinformatics tools and the functional annotations were performed. The functions of 662 HPs were successfully predicted and further classified 483 HPs as enzymes, 141 HPs were predicted to be involved in the diverse cellular mechanisms and 38 HPs may function as transporters and carriers proteins. Furthermore, 28 HPs were predicted to be virulent in nature. Amongst them, the HP P95201, HP P9WM79, HP I6WZ30, HP I6 × 9T8, HP P9WKP3, and HP P9WK89 showed the highest virulence scores. Therefore, these proteins were subjected to extensive structure analyses and dynamics of their conformations were investigated using the principles of molecular dynamics simulations, each for a 150 ns time scale. This study provides a deeper understanding of the undiscovered drug targets and the generated outputs will facilitate the process of drug design and discovery against the infection of M. tuberculosis.

[1]  Thomas L. Madden,et al.  Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. , 1997, Nucleic acids research.

[2]  Faizan Ahmad,et al.  In silico approaches for the identification of virulence candidates amongst hypothetical proteins of Mycoplasma pneumoniae 309 , 2015, Comput. Biol. Chem..

[3]  S. Ehrt,et al.  Dihydrolipoamide Acyltransferase Is Critical for Mycobacterium tuberculosis Pathogenesis , 2006, Infection and Immunity.

[4]  C. Doern,et al.  Point mutations within the streptococcal regulator of virulence (Srv) alter protein-DNA interactions and Srv function. , 2008, Microbiology.

[5]  Wei Chen,et al.  Predicting the subcellular localization of mycobacterial proteins by incorporating the optimal tripeptides into the general form of pseudo amino acid composition. , 2015, Molecular bioSystems.

[6]  E. Böttger,et al.  UvrD2 Is Essential in Mycobacterium tuberculosis, but Its Helicase Activity Is Not Required , 2011, Journal of bacteriology.

[7]  Lorena Novoa-Aponte,et al.  Mycobacterium tuberculosis P-Type ATPases: Possible Targets for Drug or Vaccine Development , 2014, BioMed research international.

[8]  A. Filloux.  Secretion Signal and Protein Targeting in Bacteria: a Biological Puzzle , 2010, Journal of bacteriology.

[9]  S. Ehlers,et al.  Lipoprotein processing is required for virulence of Mycobacterium tuberculosis † , 2004, Molecular microbiology.

[10]  M. Giffin,et al.  ald of Mycobacterium tuberculosis Encodes both the Alanine Dehydrogenase and the Putative Glycine Dehydrogenase , 2011, Journal of bacteriology.

[11]  L. Mourey,et al.  Further Insight into S-Adenosylmethionine-dependent Methyltransferases , 2006, Journal of Biological Chemistry.

[12]  G. Riccardi,et al.  Glutamine amidotransferase activity of NAD+ synthetase from Mycobacterium tuberculosis depends on an amino-terminal nitrilase domain. , 2005, Research in microbiology.

[13]  Seyed E. Hasnain,et al.  Iron-Dependent RNA-Binding Activity of Mycobacterium tuberculosis Aconitase , 2007, Journal of bacteriology.

[14]  L. Naesens,et al.  First Crystal Structures of Mycobacterium tuberculosis 6-Oxopurine Phosphoribosyltransferase: Complexes with GMP and Pyrophosphate and with Acyclic Nucleoside Phosphonates Whose Prodrugs Have Antituberculosis Activity. , 2015, Journal of medicinal chemistry.

[15]  M. Vijayan,et al.  Structure of Mycobacterium tuberculosis single-stranded DNA-binding protein. Variability in quaternary structure and its implications. , 2003, Journal of molecular biology.

[16]  L. J. Perry,et al.  Crystal structure of the apo forms of psi 55 tRNA pseudouridine synthase from Mycobacterium tuberculosis: a hinge at the base of the catalytic cleft. , 2004, The Journal of biological chemistry.

[17]  James C. Sacchettini,et al.  Structure of the Mycobacterium tuberculosis d-Alanine:d-Alanine Ligase, a Target of the Antituberculosis Drug d-Cycloserine , 2010, Antimicrobial Agents and Chemotherapy.

[18]  Burkhard Rost,et al.  Protein subcellular localization prediction using artificial intelligence technology. , 2008, Methods in molecular biology.

[19]  N. Ohara,et al.  Tetratricopeptide Repeat Protein-Associated Proteins Contribute to the Virulence of Porphyromonas gingivalis , 2010, Infection and Immunity.

[20]  Sanath H. Kumar,et al.  Biochemistry of Bacterial Multidrug Efflux Pumps , 2012, International journal of molecular sciences.

[21]  Shuangjiang Liu,et al.  Crystal Structures and Site-directed Mutagenesis of a Mycothiol-dependent Enzyme Reveal a Novel Folding and Molecular Basis for Mycothiol-mediated Maleylpyruvate Isomerization* , 2007, Journal of Biological Chemistry.

[22]  A. Krogh,et al.  Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. , 2001, Journal of molecular biology.

[23]  K. Britton,et al.  The structure and domain organization of Escherichia coli isocitrate lyase. , 2001, Acta crystallographica. Section D, Biological crystallography.

[24]  N. Prasadarao,et al.  SR-Like RNA-Binding Protein Slr1 Affects Candida albicans Filamentation and Virulence , 2013, Infection and Immunity.

[25]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[26]  J. Content,et al.  The ATP binding cassette (ABC) transport systems of Mycobacterium tuberculosis. , 2000, FEMS microbiology reviews.

[27]  G. A. Grant,et al.  Regulation of Mycobacterium tuberculosis D-3-phosphoglycerate dehydrogenase by phosphate-modulated quaternary structure dynamics and a potential role for polyphosphate in enzyme regulation. , 2014, Biochemistry.

[28]  R. Titball,et al.  Lipoproteins of Bacterial Pathogens , 2010, Infection and Immunity.

[29]  M. Carr,et al.  Mycobacterium tuberculosis RNA Polymerase-binding Protein A (RbpA) and Its Interactions with Sigma Factors* , 2013, The Journal of Biological Chemistry.

[30]  Gerrit Groenhof,et al.  GROMACS: Fast, flexible, and free , 2005, J. Comput. Chem..

[31]  J. Sacchettini,et al.  Structure, Activity, and Inhibition of the Carboxyltransferase β-Subunit of Acetyl Coenzyme A Carboxylase (AccD6) from Mycobacterium tuberculosis , 2014, Antimicrobial Agents and Chemotherapy.

[32]  M. Horwitz,et al.  All four Mycobacterium tuberculosis glnA genes encode glutamine synthetase activities but only GlnA1 is abundantly expressed and essential for bacterial homeostasis , 2005, Molecular microbiology.

[33]  R. Friedman,et al.  Bacterial luciferase is naturally destabilized in Mycobacterium tuberculosis and can be used to monitor changes in gene expression. , 2005, FEMS microbiology letters.

[34]  L. Niu,et al.  Structure of lpg0406, a carboxymuconolactone decarboxylase family protein possibly involved in antioxidative response from Legionella pneumophila , 2015, Protein science : a publication of the Protein Society.

[35]  Guo-Ping Zhao,et al.  Mycobacterial MazG is a novel NTP pyrophosphohydrolase involved in oxidative stress response. , 2010, The Journal of biological chemistry.

[36]  H. Rubin,et al.  Mycobacterium tuberculosis Type II NADH-Menaquinone Oxidoreductase Catalyzes Electron Transfer through a Two-Site Ping-Pong Mechanism and Has Two Quinone-Binding Sites , 2014, Biochemistry.

[37]  K. Poole.  Resistance to β-lactam antibiotics , 2004, Cellular and Molecular Life Sciences CMLS.

[38]  Sahadevan Raman,et al.  Transcription Regulation by the Mycobacterium tuberculosis Alternative Sigma Factor SigD and Its Role in Virulence , 2004, Journal of bacteriology.

[39]  W. Bishai,et al.  Cyclic AMP intoxication of macrophages by a Mycobacterium tuberculosis adenylate cyclase , 2008, Nature.

[40]  Teresa Quitugua,et al.  Single Nucleotide Polymorphisms in Genes Associated with Isoniazid Resistance in Mycobacterium tuberculosis , 2003, Antimicrobial Agents and Chemotherapy.

[41]  Krishna Bisetty,et al.  Current Advances in the Identification and Characterization of Putative Drug and Vaccine Targets in the Bacterial Genomes. , 2015, Current topics in medicinal chemistry.

[42]  Y. Singh,et al.  Biochemical characterization of an S-adenosyl-l-methionine-dependent methyltransferase (Rv0469) of Mycobacterium tuberculosis , 2013, Biological chemistry.

[43]  I. Smith,et al.  Mycobacterium tuberculosis Pathogenesis and Molecular Determinants of Virulence , 2003, Clinical Microbiology Reviews.