Cell wall glucomannoproteins of Saccharomyces cerevisiae mnn9

Abstract

Mannoproteins were isolated from Saccharomyces cerevisiae mnn9 mutant cell walls by laminarinase digestion and purified by affinity and anion-exchange chromatography. The purified mannoprotein fraction contained three predominant proteins with molecular masses of 300 kDa, 220 kDa and 160 kDa. These compounds were absent in an SDS extract of cell walls or in a hot-citrate extract of mnn9 cells. The carbohydrate part of the purified mannoproteins consisted of (N-acetyl)glucosamine, mannose and glucose in a molar ratio of 1:53:4. O-Glycosidically linked chains, containing 70% of the mannose, were released by mild beta-elimination. N-Glycosidically linked chains, representing 80% of the (N-acetyl)glucosamine and 20% of the mannose, were released by peptide N-glycosidase F (PNGase F) digestion. Complete degradation of protein by alkaline hydrolysis released besides the N- and O-glycosidically linked chains, another type of carbohydrate chain containing the residual (N-acetyl)glucosamine, mannose and most of the glucose in a molar ratio of 1:17:18. Glucose was beta-glycosidically linked. The results indicate that beta-glucose is linked to PNGase F-resistant N-linked chains present on cell wall mannoproteins. We propose that these chains are responsible for the linkage between mannoproteins and glucan in the cell wall.

Glucosylation of chimeric proteins in the cell wall of Saccharomyces cerevisiae

“…Cultures were grown to an OD,,, nm of 2.0. Cell walls were isolated as in [2], boiled in the presence of SDS, EDTA and /I-mercaptoethanol to obtain detergent extracts as in [7] and subsequently digested with laminarinase to obtain B-1,3glucanase extracts as in [2]. Proteins present in the culture fluid were precipitated using deoxycholate [8].…”

“…We recently demonstrated that several /I-1,3-glucanase-extractable wall proteins are covalently linked to a /I-1,6-glucan [2,3]. This raised the question whether this type of side-chain is involved in anchoring glucanaseextractable mannoproteins in the cell wall.…”

Cell Wall Proteins ofSaccharomyces cerevisiae

“…Van Rinsum et al (1991) isolated a carbohydrate chain consisting of glucosamine, mannose and~-glycosydicallylinked glucose fronl glucanase-extracted cell wall proteins in a molar ratio of 1: 17: 18. This -1 ,6-glucan chain is involved in cell wall anchorage (Kapteyn et a1., 1996).…”

Wall formation by Candida albicans yeast cells: synthesis, secretion and incorporation of two types of mannoproteins

“…Glucan and chitin in Schizophyllum cumnzune are connected by covalent bridges (Sonnenberg et al, 1982;Wessels et al, 1983), and linkages between mannoproteins and glucose molecules (glucans) have been suggested in Sacch. cerevisiae (van Rinsum et al, 1991). However, the nature of the linkages is still a matter of speculation.…”