Occurrence of a unique amino acid racemase in a basidiomycetous mushroom, Lentinus edodes

Free d-alanine was detected in a cell extract of the fruit-body of an edible basidiomycetous mushroom, Lentinus edodes (Shiitake), by means of reverse-phase high performance liquid chromatography. We also found an amino acid racemase activity in L. edodes fruit-body, and purified the enzyme. The enzyme has a molecular weight of approximately 86,000, and consists of two subunits of identical molecular weight (44,000). The optimal pH of the enzyme activity is around pH 9.5 for both d-to-l and l-to-d alanine racemization. The enzyme requires pyridoxal 5′-phosphate as a cofactor. Km and Vmax values for d-alanine were 37.3 mM and 520 nmol/min/mg, respectively; for l-alanine, they were 9.21 mM and 141 nmol/min/mg, respectively. The equilibrium constant was calculated to be 1.10, which is consistent with the theoretical value for the racemase reaction. The ability of the enzyme to catalyze the racemization of various d-amino acids was investigated. The enzyme catalyzes the racemization of d-serine (relative reaction rate, 144% of rate for d-alanine), d-alanine (100%), d-homoserine (17.1%), d-2-aminobutyrate (5.6%), d-glutamate (4.5%), and d-asparagine (3.2%). To the best of our knowledge, this is the first report of an amino acid racemase produced by a basidiomycetous mushroom.