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Directed evolution of Alcaligenes faecalis nitrilase

Paper ID Volume ID Publish Year Pages File Format Full-Text
17585 42681 2010 7 PDF Available
Title
Directed evolution of Alcaligenes faecalis nitrilase
Abstract

Alcaligenes faecalis nitrilase (NITAf) was engineered in a directed evolution approach towards increased activity at its optimal pH as well as improved fitness at low pH values. Error prone PCR in combination with recombination of beneficial mutations resulted in a variant with increased specific activity for 2-phenylpropionitrile at pH 7.5. In addition, a new nitrilase variant (pHNIT45) was developed that is catalytically active at pH values as low as pH 4.5. Within 10 min this mutant fully hydrolyzes the base labile substrate (R)-2-Cl-mandelonitrile (10 mM) to give the product (R)-2-Cl-mandelic acid (conversion 100%, ee > 99%) with full retention of enantiopurity.

Keywords
Enzyme catalysis; Nitrilase; Directed evolution; Mutation; pH stability
First Page Preview
Directed evolution of Alcaligenes faecalis nitrilase
Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 47, Issue 4, 6 September 2010, Pages 140–146
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering