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Redirecting catalysis from proteolysis to perhydrolysis in subtilisin Carlsberg

Paper ID Volume ID Publish Year Pages File Format Full-Text
23507 43444 2013 8 PDF Available
Title
Redirecting catalysis from proteolysis to perhydrolysis in subtilisin Carlsberg
Abstract

•Engineering of substrate binding pocket provided insights in protease's promiscuity.•The specificity of subtilisin Carlsberg is shifted towards peroxycarboxylic acid production.•Novel subtilisin Carlsberg variants have up to 5.5-fold increased perhydrolytic activity.

Enzyme promiscuity describes the ability of biocatalysts to catalyze conversions beyond their natural reactions. Enzyme engineering to promote side reactions is attractive for synthetic and industrial applications. For instance, a subtilisin Carlsberg protease variant (T58A/L216W) catalyzes in addition to its proteolytic activity the generation of peroxycarboxylic acids from corresponding esters in the presence of hydrogen peroxide. In the current study we used a semi-rational design approach to shift the specificity of subtilisin Carlsberg towards production of peroxycarboxylic acid. Among other identified amino acid substitutions, position Gly165 in the S1 binding pocket provided insights in subtilisin Carlsberg's promiscuity by promoting ester perhydrolysis. Catalytic constants of subtilisin Carlsberg for perhydrolysis of methyl-propionate, methyl-butyrate and methyl-pentanoate were increased up to 3.5-, 5.4- and 5.5-fold, respectively, while proteolysis was decreased up to 100-fold for N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide substrate (suc-AAPF-pNA).

Keywords
Directed evolution; Enzyme promiscuity; Perhydrolysis; Protein engineering; Subtilisin Carlsberg
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Redirecting catalysis from proteolysis to perhydrolysis in subtilisin Carlsberg
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 167, Issue 3, 10 September 2013, Pages 279–286
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
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Any Questions? feel free to contact us