Hydroxy fatty acid dehydrogenase, which is involved in polyunsaturated fatty acid saturation metabolism in Lactobacillus plantarum AKU 1009a, was cloned, expressed, purified, and characterized. The enzyme preferentially catalyzed NADH-dependent hydrogenation of oxo fatty acids over NAD(+)-dependent dehydrogenation of hydroxy fatty acids. In the dehydrogenation reaction, fatty acids with an internal hydroxy group such as 10-hydroxy-cis-12-octadecenoic acid, 12-hydroxy-cis-9-octadecenoic acid, and 13-hydroxy-cis-9-octadecenoic acid served as better substrates than those with alpha- or beta-hydroxy groups such as 3-hydroxyoctadecanoic acid or 2-hydroxyeicosanoic acid. The apparent Km value for 10-hydroxy-cis-12-octadecenoic acid (HYA) was estimated to be 38 mu M with a k(cat) of 7.6 x 10(-3) s(-1). The apparent K-m value for 10-oxo-cis-12-octadecenoic acid (KetoA) was estimated to be 1.8 mu M with a k(cat) of 5.7 x 10(-1) s(-1). In the hydrogenation reaction of KetoA, both (R)- and (S)-HYA were generated, indicating that the enzyme has low stereoselectivity. This is the first report of a dehydrogenase with a preference for fatty acids with an internal hydroxy group. (C) 2015 Elsevier B.V. All rights reserved.