THE OST4 GENE OF SACCHAROMYCES-CEREVISIAE ENCODES AN UNUSUALLY SMALL PROTEIN REQUIRED FOR NORMAL LEVELS OF OLIGOSACCHARYLTRANSFERASE ACTIVITY

Sodium vanadate is an effective drug for the enrichment of yeast mutan ts defective in glycosylation reactions that are carried out in the Go lgi complex (1). We have isolated vanadate-resistant, hygromycin B-sen sitive mutants that act at very early steps of N-linked glycosylation, occurring in the endoplasmic reticulum, Here we describe the phenotyp ic characterization of ost4, a vanadate-resistant mutant that is defec tive in oligosaccharyltransferase (OTase) activity both in vivo and in vitro. The OST4 open reading frame is unusual in that it predicts a p rotein of only 36 amino acids, We demonstrate that the OST4 gene produ ct is, in fact, an unusually small protein of approximately 3.6 kDa, p redicted to lie almost entirely in the hydrophobic environment of the membrane. Strains carrying a disruption of the OST4 gene are viable bu t grow poorly at 25 degrees C. The null mutant is inviable at 37 degre es C, demonstrating that the OST4 gene product is essential for growth at high temperatures. Deletion of the OST4 gene greatly diminishes OT ase activity but does not abolish it. These results suggest that the O ST4 gene encodes a subunit or accessory component of OTase that is ess ential at high temperature.