PREFERENTIAL TRANSFER OF TRUNCATED OLIGOSACCHARIDES TO THE FIRST SEQUON OF YEAST EXOGLUCANASE IN SACCHAROMYCES-CEREVISIAE ALG3 CELLS

In addition to the exoglucanases (Exg) secreted into the culture mediu m by wild type cells, ExgIa and ExgIb, which have oligosaccharides att ached to both potential N-glycosylation sites, Saccharomyces cerevisia e alg3 mutant secreted substantial amounts (35-44%) of underglycosylat ed and unglycosylated forms. Quantification of these forms indicated t hat no more than 78% of the available N-sites were occupied. About 50% of the transferred oligosaccharides were endo H sensitive, indicating char the lipid-linked precursor had completed its synthesis to Glc(3) -Man(9)-GlcNAc(2). The other 50% remained endo H-resistant and, accord ingly, it should be derived from the precursor oligosaccharide Man(5)- GlcNAc(2) synthesized by this mutant. A closer analysis of forms that have received two oligosaccharides (ExgIb) showed that the first sequo n was enriched in truncated residues, whereas the second one was enric hed in regular counterparts. Similarly, analysis of the individual und erglycosylated glycoforms indicated that 38% of the oligosaccharides a ttached to the second site were regular. This percentage dropped to 20 % for glycoforms carrying the oligosaccharide in the first sequon. The preferential transfer of truncated oligosaccharides to the first glyc osylation site seems to be a consequence of(1) the low percentage of t runcated lipid linked oligosaccharides that receives the glucotriose u nit, and (2) the effect of the glucotriose unit on the selection of N- sites to be glycosylated.