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Novel tools for the study of class I alpha-mannosidases: a chromogenic substrate and a substrate-analog inhibitor
T Desmet1, W Nerinckx, I Stals
1Laboratory for Biochemistry, Department of Biochemistry, Physiology, and Microbiology, Ghent University, K.L. Ledeganckstraat 35, B-9000 Ghent, Belgium.
Abstract:
The use of chromogenic substrates for evaluation of class I alpha-mannosidase is described. 2('),4(')-Dinitrophenyl-alpha-D-mannopyranoside allows rapid and sensitive assays of enzymatic activities, e.g., of heterologously expressed alpha-1,2-mannosidase from Trichoderma reesei. Interaction constants of several ligands with alpha-mannosidases from class I and II could also be determined. Furthermore, novel types of inhibitors derived from D-lyxose are presented. Methyl-alpha-D-lyxopyranosyl-(1(')-->2)-alpha-D-mannopyranoside is a potent inhibitor of the alpha-1,2-mannosidase from T. reesei (K(i)=600 microM) and since it probably spans subsites -1/+1, this disaccharide could be valuable in crystallographic studies of class I alpha-mannosidases.