Imidase, a dihydropyrimidinase-like enzyme involved in the metabolism of cyclic imides

Abstract

Imidase, which preferably hydrolyzed cyclic imides to monoamidated dicarboxylates, was purified to homogeneity from a cell-free extract of Blastobacter sp. A17p-4. Cyclic imides are known to be hydrolyzed by mammalian dihydropyrimidinases. However, imidase was quite different from known dihydropyrimidinases in structure and substrate specificity. The enzyme has a relative molecular mass of 105,000 and consists of three identical subunits. The purified enzyme showed higher activity and affinity toward cyclic imides, such as succinimide (K(m) = 0.94 mM; V(max) = 910 μmol · min-1 · mg-1), glutarimide (K(m) = 4.5 mM; V(max) = 1,000 μmol · min-1 · mg-1) and maleimide (K(m) = 0.34 mM; V(max) = 5800 μmol · min-1 · mg-1), than toward cyclic ureides, which are the substrates of dihydropyrimidinases, such as dihydrouracil and hydantoin. Sulfur-containing cyclic imides, such as 2,4-thiazolidinedione and rhodanine, were also hydrolyzed. The enzyme catalyzed the reverse reaction, cyclization, but with much lower activity and affinity. The enzyme was non-competitively inhibited by succinate, which was found to be a key compound in cyclic-imide transformation in relation with the tricarboxylic acid cycle in this bacterium, suggesting that the role of imidase is to catalyze the initial step of cyclic-imide degradation.