Saccharomyces cerevisiaeGpi3p is the UDP-GlcNAc-bind-ing andpresumed catalytic subunit of the enzyme that forms GlcNAc-phosphatidylinositol in glycosylphosphatidylinosi-tol biosynthesis. It is anessential proteinwithanEX7 Emotif that is conserved in four families of retaining glycosyl-transferases. All Gpi3ps contain a cysteine residue four residues C-terminal to EX7E. To test their importance for Gpi3p functionin vivo,Glu289 and297 in theEX7 Emotif of S. cerevisiaeGpi3p, as well as Cys301, were altered by site-specificmutagenesis, and themutant proteins tested for their ability to complement nonviable GPI3-deleted haploids
