Gpi17p does not stably interact with other subunits of glycosylphosphatidylinositol transamidase in Saccharomyces cerevisiae.
Homologues of Gpi8p, Gaa1p, Gpi16p, Gpi17p, and Cdc91p are essential components of the GPI transamidase complex that adds glycosylphosphatidylinositols (GPIs 1) to newly synthesized proteins in the ER. In mammalian cells, these five subunits remain stably associated with each other in detergent. In yeast, we find no stable stoichiometric association of Gpi17p with the Gpi8p-Gpi16p-Gaa1p core in detergent extracts. Random and site-directed mutagenesis generated mutations in several highly conserved amino acids but did not yield nonfunctional alleles of Gpi17p and a saturating screen did not yield any dominant negative alleles of Gpi17p. Moreover, Gpi8p becomes unstable when any one of the other subunits is depleted, whereas Gpi17p is slightly affected only by the depletion of Gaa1p. These data suggest that yeast Gpi17p may be able to exert its GPI anchoring function without interacting in a stable and continuous manner with the other GPI-transamidase subunits. Shutting down ER-associated and vacuolar protein degradation pathways has no effect on the levels of Gpi17p or other transamidase subunits.
Zhu
Yonghua
Y
Department of Medicine/Biochemistry, University of Fribourg, Division of Biochemistry, Chemin du Musée 5, CH-1700 Fribourg, Switzerland.
Fraering
Patrick
P
Vionnet
Christine
C
Conzelmann
Andreas
A
D000217:Acyltransferases / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D019881:Aminoacyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004795:Enzyme Stability; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006867:Hydrolases / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D011485:Protein Binding; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D013816:Thermodynamics; D014617:Vacuoles / Q000201:enzymology
D017261:Glycosylphosphatidylinositols; D021122:Protein Subunits; D000217:Acyltransferases; D019881:Aminoacyltransferases; C081007:COOH-terminal signal transamidase; C023175:transamidases; D006867:Hydrolases