화학공학소재연구정보센터
Bioresource Technology, Vol.102, No.2, 1733-1739, 2011
Purification and characterization of a novel NADH-dependent carbonyl reductase from Pichia stipitis involved in biosynthesis of optically pure ethyl (S)-4-chloro-3-hydroxybutanoate
A novel NADH-dependent dehydrogenases/reductases (SDRs) superfamily reductase (PsCRII) was isolated from Pichia stipitis. It produced ethyl (S)-4-chloro-3-hydroxybutanoate [(S)-CHBE] in greater than 99% enantiomeric excess. This enzyme was purified to homogeneity by ammonium sulfate precipitation followed by Q-Sepharose chromatography. Compared to similar known reductases producing (S)-CHBE, PsCR II was more suitable for production since the purified PsCRII preferred the inexpensive cofactor NADH to NADPH as the electron donor. Furthermore, the K(m) of PsCRII for ethyl 4-chloro-3-oxobutanoate (COBE) was 3.3 mM, and the corresponding V(max) was 224 mu mol/mg protein/min. The catalytic efficiency is the highest value ever reported for NADH-dependent reductases from yeasts that produce CHBE with high enantioselectivity. In addition, this enzyme exhibited broad substrate specificity for several beta-keto esters using NADH as the coenzyme. The properties of PsCRII with those of other carbonyl reductases from yeasts were also compared in this study. (C) 2010 Elsevier Ltd. All rights reserved.