THE GENETIC INTERACTION OF KAR2 AND WBPL MUTATIONS - DISTINCT FUNCTIONS OF BINDING-PROTEIN BIP AND N-LINKED GLYCOSYLATION IN THE PROCESSINGPATHWAY OF SECRETED PROTEINS IN SACCHAROMYCES-CEREVISIAE

The endoplasmic binding protein BiP and N-linked glycosylation are pro posed to be essential components in the processing pathway of secreted protein. In Saccharomyces cerevisiae, BiP is encoded by the KAR2 gene ; WBP1 encodes an essential component of the N-oligosaccharyltransfera se complex. wbp1 mutations result in reduced oligosaccharyltransferase activity and a temperature-sensitive phenotype. We show that a combin ation of kar2 and wbp1 mutations results in a synthetic phenotype with a strongly reduced growth rate at the permissive temperature. To inve stigate the role of N-linked glycosylation in BiP function, the proces sing of non-glycosylated carboxypeptidase was followed in different ka r2 strains at the permissive temperature. In all kar2 strains, the pro cessing of non-glycosylated carboxypeptidase Y was drastically reduced . A specific BiP/non-glycosylated carboxypeptidase Y complex was detec ted in kar2-159 and kar2-203 cells whereas the kar2-1 mutation did not result in such a complex. Our data show that BiP and N-linked glycosy lation are directly involved in the processing of secreted proteins. T he results support the hypothesis that BiP stabilizes the folding-comp etent and assembly-competent state of a polypeptide, whereas N-linked oligosaccharides are structural components required in the folding pro cess after the polypeptide is released from BiP.