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O-sialoglycoprotein endopeptidase (OSGEP) is conserved throughout almost all species like eubacteria, archaebacteria, and eukaryotes. A homolog (KAE1) of Saccharomyces cerevisiae is the essential gene for growth of the cells. Here, we designed a complementation system of KAE1- lethality using an S. cerevisiae KAE1+/- heterozygous strain. After introduction of an expression plasmid pAUR123 into the heterozygote, KAE1+ and KAE1- alleles were segregated by random spore analysis. The expression plasmid has a centromere sequence of S. cerevisiae, so is divided to each haploid cell at the same ratio. The expression of Kae1p from the plasmid could complement the lethality of KAE1 null mutation. Alanine substitution of histidine residues in a common motif of glycoprotease M22 family abolished the complementation ability of Kae1p. Therefore, the glycoprotease activity of Kae1p is essential for growth of the yeast cells. Escherichia coli and mouse OSGEP homologs also have well-conserved M22 family motifs, but did not complement the KAE1- lethality in the yeast strain. This suggests that not only the enzymatic activity of OSGEP but also the specific interaction to other proteins is critical for function of yeast OSGEP.
Research papers (publications of university or research institution)
