Optimized production, characterization and application of alkaline proteases from taxonomically assessed microbial isolates from Lonar soda lake, India

Abstract

Lonar soda lake of India was explored for isolation of the industrially important alkaline protease producers. The isolates from Lonar lake were identified based on their morphological characters, microscopic features, enzymatic profile, biochemical pattern and 16S rRNA partial gene sequences as Brachybacterium sp. LAP214, Bacillus cohnii LAP217, Bacillus pseudofirmus LAP220, Brevibacterium casei LAP223 and Halomonas venusta LAP515 and deposited in Microbial Culture Collection, NCCS, Pune (India) for public access. Production of the alkaline proteases from these isolates was optimized by adopting a one-variable-at-a-time (OVAT) approach in a submerged fermentation system in modified fermentation medium (MFM). More than 3 fold enhancement in the alkaline protease production was recorded from all these isolates after optimization of the physicochemical parameters. Molecular weights of partially purified alkaline proteases from LAP515, LAP214 and LAP223 were recorded as 20.1, 14.3 and 43 kDa respectively. Molecular weights of partially purified alkaline proteases from LAP220 and LAP217 were recorded as 29 kDa. Partially purified alkaline proteases from LAP515, LAP214, LAP220, LAP217 and LAP223 exhibited Vmax values 1000, 1290.323, 526.316, 1250 and 2000 U/mg respectively. Catalytic activities of alkaline proteases from all these isolates were partially and completely inhibited in presence of a serine protease inhibitor PMSF, at the concentrations of 1 and 10 mM respectively. The metal cations Mg2+, Ba2+, and Ca2+ enhanced the catalytic activities of these proteases. Remarkable stability of these alkaline proteases was observed in presence of selected solvents and commercial detergents. These proteases therefore can be used in various biotechnological applications.