title: 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.affiliation: Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama, 939-0398, Japan, makino@pu-toyama.ac.jp.
authors.forename: Yoshihide
authors.initials: Y
authors.lastname: Makino
authors.forename: Nobuya
authors.initials: N
authors.lastname: Itoh
mesh_terms: D019840:2-Propanol / Q000378:metabolism*; D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D012240:Rhodococcus / Q000201:enzymology* / Q000235:genetics; D012641:Selection, Genetic
chemical_list: D050505:Mutant Proteins; D000429:Alcohol Oxidoreductases; C110191:phenylacetaldehyde reductase; D019840:2-Propanol