Highly efficient synthesis of ethyl (S)-4-chloro-3-hydroxybutanoate by a novel carbonyl reductase from Yarrowia lipolytica and using mannitol or sorbitol as cosubstrate

An NADPH-dependent carbonyl reductase (YlCR2) from Yarrowia lipolytica was discovered by genome mining, overexpressed in Escherichia coli BL21 and purified to homogeneity. To efficiently synthesize ethyl (S)-4-chloro-3-hydroxybutanoate ((S)-CHBE) (99%, e.e), the highly stereoselective bioreduction of ethyl 4-chloro-3-oxobutanoate (COBE) into (S)-CHBE with the recombinant E. coli BL21/pETYlCR2 was successfully demonstrated in an n-butyl acetate-water biphasic system (1:1, v/v) with NADPH self-regeneration by substrate-coupled system using sorbitol or mannitol as co-substrate. The optimum reaction condition for the biotransformation of COBE in the biphasic system were 3000mM COBE, (1.2 or 1.3mmol/mmol COBE) mannitol or sorbitol, 0.2mM or 0.4mM NADP⁺, 0.12 or 0.14g (wet weight)/ml cell dosage, pH 5.0, 30°C; (S)-CHBE with yield of 90% and e.e of 99% was obtained after 10h reaction. Furthermore, 3000mM COBE could also be completely biotransformed after 20h without addition of expensive cofactor NADP⁺. Significantly, E. coli BL21/pETYlCR2 shows the high potential in the industrial production of (S)-CHBE.