This work describes functional and structural transitions of a novel protease isolated from Conidiobolus brefeldianus MTCC 5185 (Cprot), in detail using biophysical and bioinformatics tools. The commercial importance of Cprot in silk and leather industries made it an interesting candidate for structural investigations. Cprot possesses 8.2% α-helix, 31.1% β-sheet and 23.8% turns. The enzyme was found to be active over a wide pH range and up to 55°C. The protease was also stable in organic solvents up to 50% (v/v) concentration of alcohols and DMSO for >24h and in 2M guanidine hydrochloride for >12h. Cprot was also resistant to trypsin, chymotrypsin, proteinase K and fluorinated alcohols (5-10%). The melting temperatures observed for the native Cprot and for the enzyme treated under various stress conditions correlated well with the corresponding structural and functional transitions obtained. The structural information was supported by the homology model of its closest homologue from C. coronatus; revealing its similarity to PA clan of proteases (Proteases of mixed nucleophile, superfamily A), with His-64, Asp-113 and Ser-208 as putative catalytic triad. Three tryptophan residues in Cprot are surrounded by positively charged residues, as evident from solute quenching studies and homology model.
Keywords: Conformational transitions; Differential scanning fluorimetry; Homology model; Serine protease; Stability in organic solvents.
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