4-Methylumbelliferyl glycosides of N-acetyl 4-thiochito-oligosaccharides as fluorogenic substrates for chitodextrinase from Vibrio furnissii

Abstract

The degradation of chitin involves a diverse array of enzymes, some with overlapping substrate specificities. In order to distinguish between different types of enzymes, specific substrates containing thio-glycosidic linkages, 4-methylumbelliferyl N,N'-diacetyl-4-thio-β-chitobioside (Mu-TCB) and N,N'N''-triacetyl-4,4'-dithio-β-chitotrioside (Mu-TCT) are described. The substitution of the glycosidic oxygens (except the one that links oligosaccharide with the fluorogenic aglycon) with a sulfur atom resulted in resistance of these compounds to N-acetyl-β-hexosaminidases while they were specific substrates for the newly discovered chitodextrinase from Vibrio furnissii (Keyhani, N.O. and Roseman,S. (1996) J. Biol. Chem., 271, 33414-33424) and some bacterial chitinases. The enzyme kinetics of these 4-S-linked substrates, Mu-TCB and Mu-TCT, as well as the O-linked 4-methylumbelliferyl N,N'-diacetyl-β-chitobioside (Mu-CB) and N,N'-,N''-triacetyl-β chitotrioside (Mu-CT) with the chitodextrinase were studied and compared. The usefulness of the substrates for screening for chitodextrinase and/or chitinase activity was demonstrated.