Diversity of 4-chloroacetoacetate ethyl ester-reducing enzymes in yeasts and their application to chiral alcohol synthesis.
Enzymes which reduce 4-chloroacetoacetate ethyl ester (CAAE) to (R)- or (S)-4-chloro-3-hydroxybutanoate ethyl ester (CHBE) were investigated. Several microorganisms which can reduce CAAE with high yields were discovered. An NADPH-dependent aldehyde reductase, ARI, and an NADPH-dependent carbonyl reductase, S1, were isolated from Sporobolomyces salmonicolor and Candida magnoliae, respectively, and enzymatic synthesis of chiral CHBE was performed through the reduction of CAAE. When ARI-overproducing Escherichia coli transformant cells or C. magnoliae cells were incubated in an organic solvent-water diphasic system. CAAE was stoichiometrically converted to (R)- or (S)-CHBE (> 92% enantiomeric excess), respectively. Multiple CAAE-reducing enzymes were present in S. salmonicolor, C. magnoliae and bakers' yeast. Comparison of the primary structures of these CAAE-reducing enzymes with other protein sequences showed that CAAE-reducing enzymes are widely distributed in various protein families, and various physiological roles of these enzymes in the cell were speculated.
Kita
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K
Department of Biotechnology, Tottori University, 4-101 Koyama, Tottori 680-8552, Japan.
Kataoka
M
M
Shimizu
S
S