Microbial pathogenesis | 2021
Identification of Multi-Potent Protein Subtilisin A from halophilic bacterium Bacillus firmus VE2.
Abstract
Screening of halophiles with antimicrobial activity in saltpan soil samples from Nagapattinam district, Tamil Nadu, revealed isolate VE-2 as the most potent, identified as Bacillus firmus strain VE-2 through 16s rRNA gene sequencing. It had an optimimum growth condition (OD 3.1) and antimicrobial protein (AMP) production (450\u202fμg/mL) at 37\u202f°C, pH 8, 25% NaCl, and 36\u202fhrs incubation. SDS-PAGE analysis of the purified AMP showed the molecular weight of 36\u202fkDa. HPLC analysis of the purified AMP showed different amino acids, such as asparagines, alanine, lysine, proline, threonine, glycine, cysteine, serine, aspartic acid leucine, and valine. Further characterization and identification using FT-IR, 2D-PAGE, MALDI-TOF, and in-silico analysis showed that the isolated AMP had the highest similarity to Subtilisin-A. It showed antibacterial activity against clinical bacterial pathogens like S. aureus, S. pyogenes, C. diphtheria, E. coli, and P. aeruginosa with the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration of 2.5μg/mL and 20μg/mL and also against various fungal pathogens such as A. niger, A. flavus, C. albicans, C. tropicalis and C. parapsilosis with the MIC and minimum fungicidal concentrations of 1.25-80\u202fμg/mL. The purified AMP had excellent antioxidant potential, showed a scavenging effect against DPPH and Nitric oxide radicals, and displayed anticancer activity against HeLa cell lines with the IC50 values 53\u202fμg/mL. Hence, the purified bioactive antimicrobial peptides (AMP) could also be used in anticancer therapies.