GENETIC AND PHYSIOLOGICAL CHARACTERIZATION OF met15 MUTANTS OF SACCHAROMYCES CEREVISIAE: A SELECTIVE SYSTEM FOR FORWARD AND REVERSE MUTATIONS

Abstract

One hundred and thirty-three spontaneous and induced mutants of the met15 locus in Saccharomyces cerevisiae were characterized with respect to temperature sensitivity, osmotic remediability, interallelic complementation, and suppressibility by amber and ochre suppressors. Forty mutants are osmotic remedial; 17 of these, and no others, are also temperature-sensitive. Seven of 133 mutations are suppressible by an amber suppressor and 11 are suppressible by an ochre suppressor. Seventy percent of the mutants exhibited interallelic complementation, suggesting that the functional gene product of the met15 gene is a multimeric protein. Relative map positions of 30 met15 were estimated from the frequencies of X-ray-induced mitotic reversion of various heteroallelic diploids. All complementing nonsense mutations are located near one end of the gene in contrast to other nonsense mutations which span most of the gene, thus relating the direction of translation of the mRNA with respect to the fine-structure map. Recombination studies indicated that two of 30 mutants contained deletions of the entire met15 locus.-It was established that a variety of mutational types, including missense, nonsense, and deletions, are recovered with this unique system in which both forward and reverse mutations can be selected on the basis of methyl mercury resistance and methionine requirement of the met15 mutants.

Auxotrophs compromise cell growth and fatty acid production in Saccharomyces cerevisiae

“…[ 22 ] These strains grew normally in minimal plates supplementing the corresponding nutrition (Figure S1). While cultivating in minimal liquid media containing commonly used concentrations of 20 mg L −1 of uracil and histidine, 60 mg L −1 of leucine, [ 17 ] 20 mg L −1 of methionine and arginine, [ 24 ] 40 mg L −1 of tryptophan, [ 25 ] 30 mg L −1 of lysine, [ 26 ] and 50 mg L −1 of adenine, [ 27 ] a significant decrease of growth and FFA titers were observed ( p < 0.01). In particular, ura3 Δ, met15 Δ, leu2 Δ, arg4 Δ, and lys2 Δ strains had 13.5%–35.0% of the biomass and 23.7%–44.2% of FFA titer of the prototrophic strain (Figure 2A,B).…”

“…[22] These strains grew normally in minimal plates supplementing the corresponding nutrition (Figure S1). While cultivating in minimal liquid media containing commonly used concentrations of 20 mg L −1 of uracil and histidine, 60 mg L −1 of leucine, [17] 20 mg L −1 of methionine and arginine, [24] 40 mg L −1 of tryptophan, [25] 30 mg L −1 F I G U R E 4 Effects of using auxotrophic marker genes from α-ketoglutarate module on cell growth and FFA production. Control: prototrophic strain; his3Δ: strain ScYan8 with single deletion of gene HIS3; arg4Δ: strain ScYan13 with single deletion of gene ARG4.…”

Genetic modules for α‐factor pheromone controlled growth regulation of Saccharomyces cerevisiae

“…Met15p (synonymous with Met17p, Met25p [40][41][42]) is an O-acetylhomoserine sulfhydrolase catalyzing the terminal biosynthetic step of the yeast sulfate assimilation pathway (SAP) [43] by generating homocysteine from O-acetyl-L-homoserine and hydrogen sulfide [42,44,45]. Homocysteine is further converted either directly to methionine or indirectly to cysteine.…”