Abstract
Oligosaccharyl transferase (OT) scans and selectively glycosylates -Asn-X-Thr/Ser-motifs in nascent polypeptide chains in the endoplasmic reticulum (ER). Several groups have reported different results for the composition of this enzyme complex. In this study, using a membrane protein two-hybrid approach, the split-ubiquitin system, we show that except for Ost3p and Ost6p, all of the other subunits of OT exist as dimers or oligomers in the yeast, Saccharomyces cerevisiae. Ost3p and Ost6p behave strikingly similar in a series of genetic and biochemical assays, but clearly do not exist in the same OT complex. This observation, as well as the results in an accompanying study to analyze the composition of OT complex by blue native gel electrophoresis using a series of wild-type and mutant yeast strains strongly suggests that two isoforms of the OT complex exist in the ER, differing only in the presence of Ost3p or Ost6p. Each of these two isoforms of the OT complex specifically interacts with two structurally similar, but functionally different translocon complexes: the Sec61 and the Ssh1 translocon complexes.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Motifs
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Cell Proliferation
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Chromosomes / chemistry
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Cross-Linking Reagents / pharmacology
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Dimerization
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Electrophoresis, Polyacrylamide Gel
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Endoplasmic Reticulum / metabolism
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Epitopes / chemistry
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Galactose / metabolism
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Hexosyltransferases / chemistry*
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Hexosyltransferases / physiology*
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Immunoprecipitation
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Membrane Proteins / chemistry*
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Membrane Proteins / metabolism
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Membrane Proteins / physiology*
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Membrane Transport Proteins / chemistry*
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Models, Biological
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Mutation
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Peptides / chemistry
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Plasmids / metabolism
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Protein Binding
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Protein Isoforms
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Protein Transport
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SEC Translocation Channels
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Saccharomyces cerevisiae / enzymology*
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Saccharomyces cerevisiae Proteins / chemistry
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Saccharomyces cerevisiae Proteins / metabolism
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Saccharomyces cerevisiae Proteins / physiology*
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Two-Hybrid System Techniques
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Ubiquitin / chemistry
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beta-Galactosidase / metabolism
Substances
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Cross-Linking Reagents
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Epitopes
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Membrane Proteins
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Membrane Transport Proteins
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Peptides
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Protein Isoforms
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SEC Translocation Channels
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SSH1 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Ubiquitin
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Hexosyltransferases
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OST6 protein, S cerevisiae
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dolichyl-diphosphooligosaccharide - protein glycotransferase
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beta-Galactosidase
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Galactose