Oligosaccharide structures of the major exoglucanase secreted by Saccharomyces cerevisiae.
We have determined the structures of the N-linked carbohydrate chains, released by endo H, of exoglucanase II that are secreted by wild-type Saccharomyces cerevisiae and by the mnn1 mnn9 and mnn1 glycosylation mutants. The mnn9 mutation does not significantly affect N-linked oligosaccharides of exoglucanase II since we found almost identical structures in both mutant strains consisting of a slightly enlarged core with the basic structure shown in A (where M = mannose). Most of the molecules (77%) were phosphorylated on one of the starred mannoses (34%) or on both (43%) with a diesterified (alpha M-->P-->) or monoesterified phosphate group. In addition, some of the molecules apparently escape normal processing and retain the alpha-(1-->2)-linked mannose (italicized) and/or the three glucoses that are characteristic of the lipid-linked precursor (structure B). In the wild type, we found the same basic structure but more [formula; see text] than 90% of the molecules were modified with one to four alpha-(1-->3)-linked mannoses, which were absent in the strains bearing the mnn1 mutation (structure C). The proportion of acidic components was similar to that found in the mutants (78%), although, in this case, the monophosphorylated forms were more abundant (50%) than the diphosphorylated ones (28%). Most of the phosphate groups (69%) were diesterified by a disaccharide (alpha M-->3 alpha M-->P-->) instead of the single mannose found when the mnn1 mutation was present. In both mnn1 and wild type 10-15% of the oligosaccharides had an extra alpha-(1-->6)-linked mannose in the outer chain, a structure described in the recently isolated vrg1 mutant [Ballou, L., Hitzeman, R.A., Lewis, M. S., & Ballou, C. E. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 3209-3212].
Hernández
L M
LM
Departamento de Microbiología, Facultad de Ciencias, Universidad de Extremadura, Badajoz, Spain.
Olivero
I
I
Alvarado
E
E
Larriba
G
G
D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D002852:Chromatography, Ion Exchange; D043326:Glucan 1,3-beta-Glucosidase; D006031:Glycosylation; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000032:analysis*; D010766:Phosphorylation; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D001617:beta-Glucosidase / Q000737:chemistry* / Q000378:metabolism
D009844:Oligosaccharides; D001617:beta-Glucosidase; D043326:Glucan 1,3-beta-Glucosidase