Enterobacter sp. BK2K, screened from soil samples, can enantioselectively reduce 2-oxo-4-phenylbutanoic acid into (S)-2-hydroxy-4-phenylbutanoic acid. R-Hydroxy acid dehydrogenase (HADH) ( specific activity 62.6 U/mg) was purified from the crude extract of Enterobacter sp. BK2K, and its gene was cloned and functionally expressed in E. coli BL21. The optimal pH and temperature for the HADH activity were 6.5 and 30 degrees C, respectively. The purified enzyme catalyzes the reduction of various aromatic and aliphatic 2-oxo carboxylic acids to the corresponding ( S)-2-hydoxy carboxylic acids using NADH as cofactor. For example, the Km and k(cat)/K-m for 2-oxo-4-phenylbutaonoic acid in the presence of 2 mM NADH were 6.8 mM and 350 M-1 min(-1), respectively. For practical applications, a NADH recycle system employing the recombinant formate dehydrogenase from E. coli K12 was coupled with HADH in E. coli BL21. Using the recombinant HADH ( 110 U of 11 U/mg crude cell extract) and formate dehydrogenase ( 670 U of 67 U/mg crude cell extract) in 10 mL of 500 mM phosphate buffer ( pH 6.5), 96 mM of ( S)-phenyllactic acid (> 94% ee) and 95 mM of ( S)-2-hydroxy-4phenylbutanoic acid (> 94% ee) were produced in quantitative yields from 100 mM of phenylpyruvate and 2-oxo-4-phenylbutanoic acid.