Protein Engineering Network of Centres of Excellence of Canada, University of British Columbia, Vancouver.
The stereochemical course of the hydrolyses catalysed by two alpha-mannosidases has been determined directly by 1H NMR. Synthetic substrates were incubated with the enzymes and the anomeric configuration of the initially formed product was ascertained in each case by observation of the chemical shift of the anomeric proton at the hemiacetal centre. Both mannosidases were found to catalyse hydrolysis with retention of stereochemistry at the anomeric position. Human lysosomal alpha-mannosidase (a class II mannosidase) is a member of the glycosidase family 38 and thus has sequence similarity with several alpha-mannosidases responsible for glycoprotein biosynthesis. Jack bean alpha-mannosidase was shown to be mechanistically similar to the lysosomal enzyme and will provide a useful model system in mechanistic studies and inhibitor design.