The authors

have modelled Mce1A structure www.selleckchem.com/products/NVP-AUY922.html from find more residues 68 to 376, the N-terminal 67 residues and the C-terminal 78 residues were not modelled due to lack of homology [16]. Biopolymer module implemented in InsightII (Accelrys Inc.: San Diego, CA) was used to modify the mutated residues, from the InsightII fragment library. Using the same module, hydrogen atoms were added to both wild type and mutated protein structures at pH 7.0. The default cvff (Consistent Valence Force Field) force field [37] was applied to both the structures. Further, a series of energy minimization steps were performed on both the protein structures by InsightII/Discover (Accelrys Inc., San Diego, CA) using the following protocol: (a) In the first step of minimization, all the heavy (all non-hydrogen) atoms were constrained, the hydrogen atoms were allowed to minimize by steepest decent algorithm until the

maximum derivative (|dE/dr|) of the system was <1 kcal/(mole.Ǻ). (b) This step was followed by another steepest descent minimization with the same parameter as in step (a), but constraining the protein backbone atoms and relaxing all other atoms of the molecule. (c) In the final step, the protein molecule was minimized by conjugate gradient method with https://www.selleckchem.com/products/AG-014699.html the backbone atom fixed and allowing all other atoms relax until the maximum derivative was <0.01 kcal/(mole.Ǻ). The deviation between the two structures is evaluated by their RMSD values which could affect stability and functional

activity. Structure analysis of protein after energy minimization of protein structure was analyzed using Discovery Studio 2.5 (DS Modeling 2.5, Accelrys Inc.: IKBKE San Diego, CA). Statistical methods Statistical analysis was done by Fischer’s exact t test using Graph Pad Prism software http://​www.​graphpad.​com/​quickcalcs/​contingency1.​cfm A two-tailed p-value < 0.05 was considered statistically significant. Acknowledgements The authors thank Indian Council for Medical Research (ICMR), Govt of India for financial support. RP thank Council for Scientific and Industrial Research (CSIR), Govt of India for Senior Research Fellowship (SRF). The support from Department of Biotechnology, Govt. of India for Bioinformatics Facility (BIF) at Dr. B.R. Ambedkar Center for Biomedical Research is highly acknowledged. Electronic supplementary material Additional file 1: Overlapping primers to sequence entire mce1 and mce4 operons. (DOC 64 KB) References 1. Van Embden JD, Cave MD, Crawford JT, Dale JW, Eisenach KD, Gicquel B, Hermans P, Martin C, McAdam R, Shinnick TM: Strain identification of Mycobacterium tuberculosis by DNA fingerprinting: recommendations for a standardized methodology. J Clin Microbiol 1993, 31: 406–409.PubMed 2.