RESUMO
Enzymatic conversion of natural glycosides to their corresponding hydroxylated products using cytochromes P450 has significant advantages over synthetic chemistry and even enzyme-catalyzed glycosylation of chemicals. At present, the basic strategy for making glycosides of stilbenoid compounds is to use the glycosylation activity of enzymes, such as glycosyltransferases. Here, an efficient synthesis of a valuable (E)-astringin, a piceatannol glucoside, was developed using CYP102A1 via the highly regioselective C-3' hydroxylation of polydatin, a resveratrol glucoside. (E)-astringin is a high added value compound found in plants and wine. Benzylic hydroxylation of polydatin provides an attractive route to (E)-astringin, a catechol product. Thus far, chemical and enzymatic methods of producing (E)-astringin have not been developed. In the present study, a set of CYP102A1 mutants from Bacillus megaterium was found to catalyze regioselective hydroxylation of polydatin at the C-3' position to generate an (E)-astringin, a piceatannol glucoside.
Assuntos
Proteínas de Bactérias/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Glucosídeos/biossíntese , Glucosídeos/metabolismo , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Estilbenos/metabolismo , Substituição de Aminoácidos , Bacillus megaterium/enzimologia , Bacillus megaterium/genética , Proteínas de Bactérias/genética , Biocatálise , Biotecnologia , Sistema Enzimático do Citocromo P-450/genética , Glucosídeos/química , Hidroxilação , Cinética , Mutagênese Sítio-Dirigida , NADPH-Ferri-Hemoproteína Redutase/genética , Engenharia de Proteínas , Estilbenos/químicaRESUMO
OBJECTIVES: To find a simple enzymatic strategy for the efficient synthesis of the expensive 5'-hydroxyomeprazole sulfide, a recently identified minor human metabolite, from omeprazole sulfide, which is an inexpensive substrate. RESULTS: The practical synthetic strategy for the 5'-OH omeprazole sulfide was accomplished with a set of highly active CYP102A1 mutants, which were obtained by blue colony screening from CYP102A1 libraries with a high conversion yield. The mutant and even the wild-type enzyme of CYP102A1 catalyzed the high regioselective (98 %) C-H hydroxylation of omeprazole sulfide to 5'-OH omeprazole sulfide with a high conversion yield (85-90 %). CONCLUSIONS: A highly efficient synthesis of 5'-OH omeprazole sulfide was developed using CYP102A1 from Bacillus megaterium as a biocatalyst.
