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A NAD(P)H-dependent enoate reductase (OYE2p) from Saccharomyces cerevisiae YJM1341 was discovered by genome data mining for asymmetric reduction of (E/Z)-citral to (R)-citronellal with high enantioselectivity. This enzyme was heterologously expressed in E. coli and characterized for its biocatalytic properties. OYE2p was identified with reduction activities toward a diverse range of ɑ,β-unsaturated compounds bearing conjugated aldehyde, ketone, imide, carboxylic acid and ester. OYE2p showed the highest specific activity at 40 °C and a pH optimum at 7.0–8.0. The stability of OYE2p was rather pH-independent, and the half-life time values of the enzyme at pH 6.0–8.0 were more than 257 h. With regard to the reduction of (E)-citral and (Z)-citral, OYE2p exhibited different selectivity patterns. (E)-citral was exclusively reduced to (R)-citronellal by OYE2p in ≥ 99% ee, which was independent on pH. OYE2p produced both enantiomers of citronellal from (Z)-citral, but showed (R)-citronellal formation tendency, and the ee value of (R)-citronellal was affected by pH in the reaction system. Accordingly, the ee values for (R)-citronellal formation increased with the increasing levels of E-isomer in the (E/Z)-citral mixture as well as the increase of pH. Under the reaction conditions (30 °C and pH 8.6), using purified OYE2p as catalyst, 200 mM (E/Z)-citral (an approximately 10:9 mixture of geometric E-isomer and Z-isomer) was efficiently converted to (R)-citronellal with 88.8% ee and 87.2% yield. All these positive features demonstrate high potential of OYE2p for practical synthesis of (R)-citronellal and in asymmetric reduction of activated alkenes.