A GENETIC AND BIOCHEMICAL ANALYSIS OF MUTANTS TO TRYPTOPHAN INDEPENDENCE IN SACCHAROMCYES

L W Parks, H C Douglas
1957 Genetics  
T has been shown that reversion to growth factor independence in auxotrophic I strains of Neurospora may result from mutation of the defective locus itself, or mutation at a different locus (WAGNER and MITCHELL 1955) . Mutant genes of the latter type have been designated as suppressors, and biochemical studies of suppressed mutants have indicated that the metabolic pathway concerned may be different from or similar to that of wild type (LEIN and LEIN 1952; YANOFSKY 1952a) . Suggestive evidence
more » ... uggestive evidence that at least two classes of mutants to tryptophan independence occurred in Saccharomyces was obtained when it was observed that two kinds of clones developed when auxotrophic stocks were plated on tryptophanless medium. The genetic and biochemical properties of these mutants are the subject of this paper. MATERIALS AND METHODS The tryptophan requiring haploids as well as the tester stocks were obtained from DRS. D. C. HAWTHORNE and H. L. ROMAN. Clones containing the genes tr-1 and tr-2 were obtained orginally from DRS. SEYMOUR POMPER (POMPER and BURKHOLDER 1949) and CAROLINE RAUT. The results of crosses indicate that tr-1 and tr-2 are not allelic and that tr-1 is closely linked to its centromere (HAWTHORNE 1955a, b). Indole or tryptophan can fulfill the tryptophan requirement of both tr-1 and tr-2 clones, while tr-2 can use anthranilic acid as well. Clones with tr-1 accumulate both anthranilic acid and N-fructosyl-anthranilic acid in their growth medium (PARKS and DOUGLAS 1956) which suggests that tr-1 clones are blocked in their conversion of these compounds to indole. Stocks that are doubly recessive (tr-I, tr-2) accumulate neither of these compounds, indicating that tr-2 controls a step in tryptophan biosynthesis which occurs prior to the formation of anthranilic acid. Indole synthesis from anthranilic acid was determined by using 0.25 gm dry weight of a 48-hour culture of the test organism grown in complete medium. Each assay flask contained 5 ml of yeast cells in water, 500 pg anthranilic acid, 150 pg hydroxylamine (YANOFSKY 1952b), and 4.4 ml of M/10 phosphate buffer at pH 7.0 to give a total volume of 10 ml. The reaction mixtures were incubated a t room temperature on a rotary shaker for five hours. Indole was extracted in toluene and determined by
doi:10.1093/genetics/42.3.283 fatcat:zan65hd5mvh33oxbwuas6iokhy