FUNCTIONAL-ANALYSIS OF MUTATIONS IN THE PIS GENE, WHICH ENCODES SACCHAROMYCES-CEREVISIAE PHOSPHATIDYLINOSITOL SYNTHASE

The PIS gene for an enzyme phosphatidylinositol synthase having an inc reased K-m for myo-inositol, was isolated from Saccharomyces cerevisia e. The mutant PIS gene contained a CAA codon at position 114 instead o f the CAC codon observed in the wild-type gene, resulting in alteratio n of the amino acid from His to Gln. Oligonucleotide mediated site-dir ected mutagenesis of PIS at codon 114 revealed that mutant genes with codons for Ala, Thr and Leu could support yeast eel growth in vivo, bu t those, for Asp, Lys and Tyr could not. All mutant enzymes when expre ssed in Escherichia coli showed greatly reduced in vitro activity.