Most microbial D-hydantoinases have been reported to have catalytic properties similar to those of mammalian dihydropyrimidinases. Comparison of the primary structures of microbial D-hydantoinases with mammalian dihydropyrimidinases revealed that the amino acid homology is about 37% and functionally important residues are rigidly conserved at identical positions. Interestingly, however, the C-terminal regions mere found to be completely mismatched with each other. In order to investigate the possible role of the C-terminal regions, we deleted the C-terminal regions of the D-hydantoinases from two thermophilic Bacilli and compared the catalytic and structural properties of the mutant enzymes with those of wildtype enzymes. As a result, the C-terminal region was found not to be essential for catalysis, but it does affect the oligomeric structure of the enzyme. (C) 1998 Academic Press.