Structure of the Linkage Region between the Polysaccharide and Protein Parts of Saccharomyces cerevisiae Mannan

Abstract

An oligosaccharide fragment corresponding to the linkage region between the polysaccharide and protein parts of Saccharomyces cerevisiae X2180 yeast mannan has been isolated. This was accomplished by the combined action of a bacterial endo-α(1→6)mannanase and an endo-β-N-acetylglucosaminidase on the mutant mannan from S. cerevisiae X2180-1A-5 that has an unbranched polysaccharide outer chain. The mannanase removed the unbranched mannan outer chain and the glucosaminidase apparently split a di-N-acetylchitobiose unit at the point of attachment of the polysaccharide to asparagine in the protein part. This order of degradation is assumed from the facts that wild type mannan with a branched outer chain was not attacked whereas ovalbumin was degraded with the release of the expected oligosaccharide fragment terminated by N-acetylglucosamine at the reducing end. Thus, the glucosaminidase was able to act only on a partially degraded mannan chain. The linkage fragment was composed of about 12 mannose units and a single N-acetylglucosamine which was found at the reducing end. The attachment of mannose to N-acetylglucosamine was by a β(1→4) linkage as in other glycoproteins. Acetolysis of the oligosaccharide yielded mannose, mannobiose αMan(1→2)Man and αMan(1→3)Man, mannotriose with both 1→2 and 1→3 linkages, and mannotetraose, probably αMan(1→3)αMan(1→2)αMan(1→3)Man. These fragments were interlinked by α(1→6) bonds, and the oligosaccharide was attached to the N-acetylglucosamine. Another acetolysis fragment isolated, αMan(1→3)αMan(1→2)αMan(1→3)βMan(1→4)GNAc, is common to other glycoproteins. The observation that this kind of branched inner core structure is made in a yeast mutant that forms an unbranched outer chain demonstrates that the formation of these two parts of the molecule is regulated in part by different enzymes and may serve different functions in the translocation of the mannan proteins and their organization in the cell wall.