Effects of GSH1 and GSH2 Gene Mutation on Glutathione Synthetases Activity of Saccharomyces cerevisiae

摘要

In this paper, three mutants from wild Saccharomyces cerevisiae HBU2.558, called U2.558, UN2.558, and UNA2.558, were screened by UV, sodium nitrite, Atmospheric and room temperature plasma, respectively. Glutathione production of the three mutants increased by 41.86, 72.09 and 56.76%, respectively. We detected the activity of glutathione synthetases and found that its activity was improved. Amino acid sequences of three mutant colonies were compared with HBU2.558. Four mutants: Leu51 -> Pro51 (L51P), Glu62 -> Val62 (E62V), Ala332 -> Glu332 (A332E) and Ser653 -> Gly653 (S653G) were found in the analysis of gamma-glutamylcysteine ligase. L51 is located adjacently to the two active sites of GCL/E/Mg2+/ADP complex in the overall GCL structure. L51P mutant spread distortion on the beta-sheet due to the fact that the phi was changed from -50.4A degrees to -40.2A degrees. A mutant Leu54 -> Pro54 (L54P) was found in the analysis of glutathione synthetase, and L54 was an amino acid located between an alpha-helix and a beta-sheet. The results confirm that introduction of proline located at the middle of the beta-sheet or at the N- or C-terminal between alpha-helix and beta-sheet or, i.e., L51P and L54P, changed the phi, rigidity, hydrophobicity and conformational entropy, thus increased protein stability and improved the enzyme activity.