Molecular Cloning, Sequence and Structural Analysis of Dehairing Mn2+ Dependent Alkaline Serine Protease (MASPT) of Bacillus pumilus TMS55

Kalibulla      Syed Ibrahim; Jeyaraj      Muniyandi; Shunmugiah      Karutha Pandian

Abstract

Leather industries release a large amount of pollution-causing chemicals which creates one of the major industrial pollutions. The development of enzyme based processes as a potent alternative to pollution-causing chemicals is useful to overcome this issue. Proteases are enzymes which have extensive applications in leather processing and in several bioremediation processes due to their high alkaline protease activity and dehairing efficacy. In the present study, we report cloning, characterization of a Mn2+ dependent alkaline serine protease gene (MASPT) of Bacillus pumilus TMS55. The gene encoding the protease from B. pumilus TMS55 was cloned and its nucleotide sequence was determined. This gene has an open reading frame (ORF) of 1,149 bp that encodes a polypeptide of 383 amino acid residues. Our analysis showed that this polypeptide is composed of 29 residues N-terminal signal peptide, a propeptide of 79 residues and a mature protein of 275 amino acids. We performed bioinformatics analysis to compare MASPT enzyme with other proteases. Homology modeling was employed to model three dimensional structure for MASPT. Structural analysis showed that MASPT structure is composed of nine α-helices and nine β-strands. It has 3 catalytic residues and 14 metal binding residues. Docking analysis showed that residues S223, A260, N263, T328 and S329 interact with Mn2+. This study allows initial inferences about the structure of the protease and will allow the rational design of its derivatives for structure-function studies and also for further improvement of the enzyme.

Keywords: Bacillus pumilus, dehairing, alkaline serine protease, homology modeling, MASPT, ORF, TMS55, rRNA gene amplification, PCR, MODELLER, HBOND, lambda PR, NMITLI, CSIRBacillus pumilus, dehairing, alkaline serine protease, homology modeling, MASPT, ORF, TMS55, rRNA gene amplification, PCR, MODELLER, HBOND, lambda PR, NMITLI, CSIR