A novel enzyme, N-acylamino acid racemase, was purified to homogeneity from Streptomyces atratus Y-53 and characterized. This enzyme catalyzes the interconversion of optically active N-acylamino acids. The relative molecular mass (M(r)) of the enzyme was estimated to be about 41000 and 244000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration, respectively, indicating that the enzyme is composed of six subunits with an equal M(r). The enzyme showed a broad substrate specificity toward N-acylamino acids, such as N-acetylmethionine, N-chloroacetylphenylalanine and N-chloroacetylvaline. The apparent Michaelis constant (K-m) values for N-acetyl-D-methionine and N-acetyl-D-methionine were calculated to be 15.2 and 5.6 mM, respectively. Enzyme activity was markedly enhanced by divalent metal ions, such as Co2+, Mg2+ and Mn2+, and was inhibited by metal-chelating reagent, indicating that the enzyme is a metalloenzyme. We propose to name the enzyme N-acylamino acid racemase (acylamino acid racemase).