Glycosylphosphatidylinositol membrane anchors in Saccharomyces cerevisiae: absence of ceramides from complete precursor glycolipids.
Glycosylphosphatidylinositol (GPI) anchoring of membrane proteins occurs through two distinct steps, namely the assembly of a precursor glycolipid and its subsequent transfer onto newly synthesized proteins. To analyze the structure of the yeast precursor glycolipid we made use of the pmi40 mutant that incorporates very high amounts of [3H]mannose. Two very polar [3H]mannose-labeled glycolipids named CP1 and CP2 qualified as GPI precursor lipids since their carbohydrate head group, Man alpha 1,2(X-->PO4-->6)Man alpha 1,2Man alpha 1,6Man alpha-GlcN-inositol (with X most likely being ethanolamine) comprises the core structure which is common to all GPI anchors described so far. CP1 predominates in cells grown at 24 degrees C whereas CP2 is induced by stress conditions. The apparent structural identity of the head groups suggests that CP1 and CP2 contain different lipid moieties. The lipid moieties of both CP1 and CP2 can be removed by mild alkaline hydrolysis although the protein-bound GPI anchors made by the pmi40 cells under identical labeling conditions contain mild base resistant ceramides. These findings imply that the ceramide moiety found on the majority of yeast GPI anchored proteins is added through a lipid remodeling step that occurs after the addition of the GPI precursor glycolipids to proteins.
Sipos
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Institute of Biochemistry, University of Fribourg, Switzerland.
Puoti
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Conzelmann
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D000468:Alkalies; D002240:Carbohydrate Sequence; D002518:Ceramides / Q000032:analysis*; D003513:Cycloheximide / Q000494:pharmacology; D006017:Glycolipids / Q000737:chemistry* / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000737:chemistry* / Q000378:metabolism; D006868:Hydrolysis; D008358:Mannose / Q000378:metabolism; D008969:Molecular Sequence Data; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology
D000468:Alkalies; D002518:Ceramides; D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D014415:Tunicamycin; D003513:Cycloheximide; D010727:Phosphoric Diester Hydrolases; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D008358:Mannose