Ubiquitination Process on Regulator of SPS Pathway in Saccharomyces cerevisiae

Abstract:

The ubiquitination mechanism of key regulator Stp1p was investigated in Saccharomyces cerevisiae. The ubiquitination detection vector was constructed based on the BiFC（Bimolecular fluorescence complementation） technology to assay the ubiquitination process of Stp1p. The site-directed mutagenesis on the potential ubiquitination sites were performed to verify the effect on its ubiquitination regulation and the nitrogen utilization. The fluorescence levels of mutant strains were down-regulated compared to the wild type strain. Furthermore，the triple mutant Stp1K49，129，113R achieved the lowest level among the all of the strains. The results indicated that the process of ubiquitination was significantly repressed by removing the potential ubiquitination residues. In addition，the amino acid utilization assay implied the ubiquitination sites played a vital role on its ubiquitination process. The results presented here demonstrated that the ubiquitination process was invovled in the regulation of Stp1p by site-directed mutagenesis of potential ubiquitination sites and thus impact the nitrogen utilization process.