Characterization and mutation analysis of a halotolerant serine protease from a new isolate of Bacillus subtilis

• Main Authors: Shinji Takenaka, Jyun Yoshinami, Ampin Kuntiya, Charin Techapun, Noppol Leksawasdi, Phisit Seesuriyachan, Thanongsak Chaiyaso, Masanori Watanabe, Kosei Tanaka, Ken ichi Yoshida
• Format: Journal
• Published: 2018
• Subjects: Biochemistry, Genetics and Molecular Biology
• Online Access: https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=85031898530&origin=inward; http://cmuir.cmu.ac.th/jspui/handle/6653943832/58337
• Institution: Chiang Mai University

© 2017, Springer Science+Business Media B.V. Objectives: A bacterial halotolerant enzyme was characterized to understand the molecular mechanism of salt adaptation and to explore its protein engineering potential. Results: Halotolerant serine protease (Apr_No16) from a newly isolated Bacillus subtilis strain no. 16 was characterized. Multiple alignments with previously reported non-halotolerant proteases, including subtilisin Carlsberg, indicated that Apr_No16 has eight acidic or polar amino acid residues that are replaced by nonpolar amino acids in non-halotolerant proteases. Those residues were hypothesized to be one of the primary contributors to salt adaptation. An eightfold mutant substituted with Ala residues exhibited 1.2- and 1.8-fold greater halotolerance at 12.5% (w/v) NaCl than Apr_No16 and Carlsberg, respectively. Amino acid substitution notably shifted the theoretical pI of the eightfold mutant, from 6.33 to 9.23, compared with Apr_No16. The resulting protein better tolerated high salt conditions. Conclusions: Changing the pI of a bacterial serine protease may be an effective strategy to improve the enzyme’s halotolerance.