Development of an improved phenylacetaldehyde reductase mutant by an efficient selection procedure.

Abstract

Chiral alcohols are valuable as diverse chemicals and synthetic intermediate materials. Phenylacetaldehyde reductase (PAR) is an enzyme that converts a wide variety of ketones into chiral alcohols with high optical purity. When an alcohol such as 2-propanol is used as a hydrogen donor, PAR itself will also mediate the regeneration of the coenzyme NADH in situ. Perceiving a capacity for improvement, we sought to develop a PAR that is able to convert higher concentrations of substrates in the presence of high concentrations of 2-propanol. The selection procedure for mutants was re-examined and a procedure able to select an effective amino acid substitution was established. Two advantageous amino acid substitutions were successfully selected using the procedure. When high-concentration substrate conversion reaction was subjected with a mutant that integrated both the two amino acid substitutions, near-complete conversions of m-chlorophenacyl chloride (m-CPC) (2.1 mmol/ml) and ethyl 4-chloro-3-oxobutanoate (ECOB) (1.9 mmol/ml) were achieved.

• Authors: Yoshihide Makino, Nobuya Itoh
• Journal: Applied microbiology and biotechnology (Appl Microbiol Biotechnol) Vol. 98 Issue 10 Pg. 4437-43 (May 2014) ISSN: 1432-0614 [Electronic] Germany
• PMID: 24352730 (Publication Type: Journal Article)

Chemical References

• Mutant Proteins
• Alcohol Oxidoreductases
• phenylacetaldehyde reductase
• 2-Propanol

Topics

• 2-Propanol (metabolism)
• Alcohol Oxidoreductases (genetics, metabolism)
• Amino Acid Substitution
• Mutant Proteins (genetics, metabolism)
• Rhodococcus (enzymology, genetics)
• Selection, Genetic