Graduate Thesis Or Dissertation
 

Study of sulfite mutants of Saccharomyces cerevisiae

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  • Sulfite mutants representing five complementation groups, previously derived from an ethyl methanesulfonate-treated haploid strain of Saccharomyces cerevisiae were studied. Although the wildtype S. cerevisiae strain used (isogenic to X2180-1 A) had a basal tolerance for sulfite (7 μM free H₂SO₃), the sensitive and resistant mutants were found to tolerate less than 3 to 5.5, or greater than 19 μM free H₂SO₃, respectively. No apparent correlation was found between the response to sulfite and generation time in rich (YEPD) or minimal media. Resistant mutant 11-1 had an extended lag phase relative to wildtype. Mutant and wildtype proteins were labeled with ³⁵S-methionine to determine differences in banding patterns due to sulfite-specific induction or disappearance of polypeptides. No obvious differences following SDS-PAGE and autoradiography were observed upon induction with 0.213 μM free H₂SO₃. No consistent correlations were found between the sulfite phenotypes and responses to other reducing agents. Sensitive mutant 35-2 appeared to be three to ten times more sensitive to dithiothreitol than wildtype and sensitive mutant 47-9 was three to four times more sensitive to sodium nitrite and three to seven times more sensitive to sodium thiosulfate than wildtype. Log phase cells of sensitive mutant 33-2 were found to have significantly less glutathione than wildtype. Wildtype contained 62.6 nmol min⁻¹ mg protein⁻¹ (62.6 mU mg protein⁻¹) glutathione reductase (GR) and 2.78 nmol min⁻¹ mg protein⁻¹ (2.78 mU mg protein⁻¹) glutathione S-transferase (GST). Log phase cells of one resistant mutant showed a significantly higher level of GR than wildtype, 135%. The resistant mutants as well as some of the sensitive mutants had reduced GST levels. Survival rates of the mutants in buffer in the presence of sulfite did not correlate with their sensitive or resistant phenotypes, suggesting that survival and growth in the presence of sulfite are not necessarily related functions. Relative to wildtype, survival upon prolonged storage at 4°C was markedly reduced for two of the four sensitive mutants, one of which was 33-2, and was enhanced for one resistant and another sensitive mutant.
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