RESUMO
The human fecal anaerobe Eubacterium ramulus is capable of degrading various flavonoids, including the flavone naringenin. The first step in the proposed degradation pathway is the isomerization of naringenin to the corresponding chalcone. Cell-free extracts of E. ramulus displayed chalcone isomerase activity. The enzyme from E. ramulus was purified to homogeneity. Its apparent molecular mass was estimated to be 136 and 129 kDa according to gel filtration and native polyacrylamide gel electrophoresis, respectively. Chalcone isomerase is composed of one type of subunit of 30 kDa. The purified enzyme catalyzed the isomerization of naringenin chalcone, isoliquiritigenin, and butein, three chalcones that differ in their hydroxylation pattern. N-bromosuccinimide, but also naringenin and phloretin, inhibited the purified enzyme considerably. This is the first report on a bacterial chalcone isomerase. The physiological function of the purified enzyme is unclear, but an involvement in the conversion of the flavanone naringenin to the chalcone is proposed.
Assuntos
Chalcona/análogos & derivados , Chalcona/metabolismo , Eubacterium/enzimologia , Flavanonas/metabolismo , Liases Intramoleculares/isolamento & purificação , Liases Intramoleculares/metabolismo , Bromosuccinimida/farmacologia , Chalcona/química , Chalconas , Cromatografia em Gel , Eletroforese em Gel de Poliacrilamida , Inibidores Enzimáticos/farmacologia , Flavanonas/química , Flavanonas/farmacologia , Flavonoides/metabolismo , Liases Intramoleculares/química , Isomerismo , Peso Molecular , Floretina/química , Floretina/farmacologia , Subunidades Proteicas/análise , Especificidade por SubstratoRESUMO
An NADH oxidase from the strictly anaerobic Eubacterium ramuluswas purified to homogeneity. The enzyme is composed of two types of subunits with molecular masses of 40 and 30 kDa. The molecular mass of the native enzyme is 450 kDa according to gel filtration and PAGE analysis. Six to eight mol of FAD were found per mol of native enzyme. The NADH-specific enzyme was inhibited by N-bromosuccinimide and sulfhydryl reagents such as N-ethylmaleimide, CuCl(2) or ZnCl(2). The physiological function of the purified enzyme is unclear, but the demonstration of NADH-dependent O(2)-consumption suggests that it plays a role in the scavenging of oxygen.
