Cloning, expression and characterization of an alkaline protease from Bacillus sp. N1

Abstract:

[Background] Proteases are widely used in the leather industry because the enzymatic dehairing approaches are environment-friendly. However, the instability to chemical reagents and the collagen degradation activity of proteases limit their industrial applications. [Objective] An alkaline protease gene from genome of Bacillus sp. N1 was cloned and expressed in Escherichia coli. The enzymatic properties of the recombinant protease were characterized and its application in dehairing process was discussed. [Methods] Genomic library was constructed to clone the protease gene aprG. The recombinant strain Escherichia coli BL21(DE3) pLysS/pET-28a-aprG was constructed for the protease expression with the induction of isopropyl β-D-thiogalactoside (IPTG). Characterization of the protease was performed through the forint phenol chromogenic method. The dehairing activity to the sheep and rabbit skin as well as feather of the protease was investigated. [Results] An alkaline protease gene aprG was cloned and expressed in E. coli expression system. The characterization of AprG showed that the optimum temperature and pH was 50 °C and 10.0, respectively. The metal ions showed soft influence on the AprG activity. Moreover, the protease AprG exhibited outstanding tolerance to surfactants, oxidants and reducing agents. The investigation on substrate specificity demonstrated that the protease displayed low hydrolysis ability toward collagen. The application study showed that AprG demonstrated effective dehairing on sheep and rabbit skin, and also showed high feather degradation activity. [Conclusion] The protease AprG has significant application potential in the leather industry.