CLONING AND SEQUENCING OF THE GMP SYNTHETASE-ENCODING GENE OF SACCHAROMYCES-CEREVISIAE

We have localised, within a Saccharomyces cerevisiae genomic fragment, the GUA1 gene whose amplification leads to the accumulation of severa l polypeptides on the two-dimensional (2-D) map of yeast proteins. Com parison of the sequence of the putative GUA1 protein with a data libra ry shows a strong similarity with Escherichia coli, Bacillus subtilis and Dictyostelium discoideum GMP synthetases (GMPS) and other glutamin e amidotransferases. The fact that disruption of the chromosomal copy of the gene leads to guanine auxotrophy, that the gua1::URA3 disruptio n does not complement an independently obtained gua1-3 mutation defici ent in GMPS and that GUA1 complements this latter mutation, confirms t he identification of the cloned gene as GUA1 encoding the S. cerevisia e GMPS. Finally, using microsequencing, we have identified one of the polypeptides, which is overproduced in response to GUA1 amplification, as corresponding to GUA1.