Enhancement of the catalytic efficiency and thermostability of Stenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF.

ABSTRACT: In this study, we enhanced the catalytic efficiency and thermostability of keratinase KerSMD by replacing its N/C-terminal domains with those from a homologous protease, KerSMF, to degrade feather waste. Replacement of the N-terminal domain generated a mutant protein with more than twofold increased catalytic activity towards casein. Replacement of the C-terminal domain obviously improved keratinolytic activity and increased the k(cat)/K(m) value on a synthetic peptide, succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, by 54.5%. Replacement of both the N- and C-terminal domains generated a more stable mutant protein, with a Tm value of 64.60?±?0.65°C and a half-life of 244.6?±?2?min at 60°C, while deletion of the C-terminal domain from KerSMD or KerSMF resulted in mutant proteins exhibiting high activity under mesophilic conditions. These findings indicate that the pre-peptidase C-terminal domain and N-propeptide are not only important for substrate specificity, correct folding and thermostability but also support the ability of the enzyme to convert feather waste into feed additives.

SUBMITTER: Fang Z 

PROVIDER: S-EPMC4720410 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature