A genomic library of Bifidobacterium bifidum constructed in Escherichia coli was screened for the ability to hydrolyze the ␣-(132) linkage of 2-fucosyllactose, and a gene encoding 1,2-␣-L-fucosidase (AfcA) was isolated. The afcA gene was found to comprise 1,959 amino acid residues with a predicted molecular mass of 205 kDa and containing a signal peptide and a membrane anchor at the N and C termini, respectively. A domain responsible for fucosidase activity (the Fuc domain; amino acid residues 577 to 1474) was localized by deletion analysis and then purified as a hexahistidine-tagged protein. The recombinant Fuc domain specifically hydrolyzed the terminal ␣-(132)-fucosidic linkages of various oligosaccharides and a sugar chain of a glycoprotein. The stereochemical course of the hydrolysis of 2-fucosyllactose was determined to be inversion by using 1 H nuclear magnetic resonance. The primary structure of the Fuc domain exhibited no similarity to those of any glycoside hydrolases (GHs) but showed high similarity to those of several hypothetical proteins in a database. Thus, it was revealed that the AfcA protein constitutes a novel inverting GH family (GH family 95).␣-L-Fucosyl residues are frequently found at the nonreducing termini of various glycoconjugates, including blood group substances, milk oligosaccharides, gastric and submaxillary mucins, and serum glycoproteins (30, 37). The results of recent studies indicate that such terminal fucosyl residues attached by

Rhodococcus rubropertinctus N82 possesses unique regiospecific hydroxylation activity in biotransformation of compounds. In this study, the ability of whole cells of the strain R. rubropertinctus N82 in biotransformation was studied. The hydroxylation activity resulted in transforming 6,7-dihydro-4H-thieno[3,2-c]-pyridine-5-carboxylic acid tert-butyl ester (LS1) into 2-hydroxy-6,7-dihydro-4H-thieno[3,2-c]-pyridine-5-carboxylic acid tert-butyl ester (LP1), a pharmaceutical intermediate. By optimizing conditions for the hydroxylating biotransformation using whole cells of R. rubropertinctus N82 as biocatalyst, 3.3 mM LP1 was successfully produced from 4 mM LS1 with a molar yield of 83%. Thus, effective method was newly developed to produce LP1, which is a synthetic intermediate of a platelet inhibitor active pharmaceutical ingredient drug, prasugrel.