Our objective is to understand the role of glycoproteins in membranes and secretions of eukaryotes, and the factors which control the specific carbohydrate structures formed in glycoproteins since these are involved in intercellular and intracellular recognition phenomena. We are studying the biosynthesis of N-linked oligosaccharides in Saccharomyces cerevisiae which begins as in all eukaryotes with the transfer of a preformed glucose-containing high mannose oligosaccharide from a dolichyl pyrophosphate intermediate to protein. This oligosaccharide then undergoes processing which requires the concerted action of glycosidases and glycosyltransferases to obtain the completed mannoproteins. We plan to purify and characterize the Alpha-mannosidase(s) involved in processing in this organism, to determine whether one or more of these enzymes is required and what specificity is associated with each Alpha-mannosidase. We also plan to study the importance of Alpha-mannosidase on subsequent processing in yeast by studying the effect of specific inhibitors of Alpha-mannosidase(s) and by obtaining mutants lacking this activity. Such mutants will be useful to eventually clone the gene for processing Alpha-mannosidase(s). These studies will contribute to understanding the control of glycosylation in eukaryotes.