A new yeast mutation in the glucosylation steps of the asparagine-linked glycosylation pathway. Formation of a novel asparagine-linked oligosaccharide containing two glucose residues

J Biol Chem. 1986 Nov 25;261(33):15582-90.


We have isolated and characterized a new yeast mutation in the glucosylation steps of lipid-linked oligosaccharide biosynthesis, alg8-1. Cells carrying the alg8-1 mutation accumulate Glc1Man9GlcNAc2-lipid both in vivo and in vitro. We present evidence showing that the alg8-1 mutation blocks addition of the second alpha 1,3-linked glucose. alg8-1 cells transfer Glc1Man9GlcNAc2 to protein instead of the wild type oligosaccharide, Glc3Man9GlcNAc2. Pulse-chase studies indicate that the Glc1Man9GlcNAc2 transferred is processed more slowly than the wild type oligosaccharide. The yeast mutation gls1-1 lacks glucosidase I activity (Esmon, B., Esmon, P.C., and Schekman, R. (1984) J. Biol. Chem. 259, 10322-10327), the enzyme responsible for removing the alpha 1,2-linked glucose residues from protein-linked oligosaccharides. We demonstrate that gls1-1 cells contain glucosidase II activity (which removes alpha 1,3-linked glucose residues) and have constructed the alg8-1 gls1-1 haploid double mutant. The Glc1Man9GlcNAc2 oligosaccharide was trimmed normally in these cells, demonstrating that the alg8-1 oligosaccharide contained an alpha 1,3-linked glucose residue. A novel Glc2 compound was probably produced by the action of the biosynthetic enzyme that normally adds the alpha 1,2-linked glucose to lipid-linked Glc2Man9GlcNAc2. This enzyme may be able to slowly add alpha 1,2-linked glucose residue to protein-bound Glc1Man9GlcNAc2. The relevance of these findings to similar observations in other systems where glucose residues are added to asparagine-linked oligosaccharides and the possible significance of the reduced rate of oligosaccharide trimming in the alg mutants are discussed.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Asparagine / metabolism*
  • Carbohydrate Conformation
  • Dolichol Monophosphate Mannose / biosynthesis
  • Glucose / metabolism*
  • Glycoproteins / biosynthesis*
  • Glycosylation
  • Kinetics
  • Lipid Metabolism
  • Mannose / metabolism
  • Mutation*
  • Oligosaccharides / biosynthesis*
  • Oligosaccharides / genetics
  • Polyisoprenyl Phosphate Monosaccharides / biosynthesis
  • Saccharomyces cerevisiae / genetics*


  • Glycoproteins
  • Oligosaccharides
  • Polyisoprenyl Phosphate Monosaccharides
  • Dolichol Monophosphate Mannose
  • dolichol-D-glucosylmonophosphate
  • Asparagine
  • Glucose
  • Mannose