Effect Of Deficiency Of Vacuolar Proteinase A In Saccharomvces Cerevisiae On Its Glycometabolism And Stress Resistance Property

Saccharomyces cerevisiae proteinase A (PrA), encoding by PEP4gene, is a member of the aspartic proteinase superfamily, which belongs to PGA family. PrA is essential to S. cerevisiae vacuolar proteolytic system during nutritional stress, sporulation and vegetative growth conditions. PrA has been implicated in the activities of other hydrolases, including proteinase B (PrB), carboxypeptidase(CPY) and aminopeptidase I. In recent years, many researches identified the unique biological function of PrA in S. cerevisiae and gradually realized the important role of PrA in S. cerevisiae. Though the understanding of structure and functions of PrA in S. cerevisiae is well known, its regulatory effect on cell physiology and metabolism is still understood. Based on this, we carried out this study and further research in the effect of deficiency of vacuolar proteinase A in Saccharomyces cerevisiae on its glycometabolism and stress resistance property was studied.This paper mainly focused on the following contents including:the influence caused by the knock out of proteinase A on the key enzymes specific activity in glycometabolism, influence on the key enzymes gene expression, influence on the related protein expression, influence on the stress resistance property under the circumstance of low-nitrogen, oxidation and radiation conditions. And the results showed that the knock-out of PEP4caused the up-regulation of the key enzymes specific activity in glycometabolism, the six key enzymes are HK, PFK, PK, CS, ICD and a-KGDHC; the knock-out of PEP4raised the gene expression of CS and a-KGDHC, depressed the gene expression of HK, PFK, PK and ICD, and may probably has an effect on the post-translation and modification of these enzymes. In addition, the knock-out of PEP4caused significant down-regulation of Fbalp, Eno2p and Pdclp, further RT-qPCR results showed that Pdclp, is the most influenced enzyme among these enzymes. Combined with the overview of haploid S. cerevisiae glycolytic metabolic processes, it can be inferred that the absence of proteinase A may probably influence the whole cell biochemical metabolism of S. cerevisiae. Upon nitrogen starvation circumstances, the knock-out of PEP4gene caused the cell autolysis and dissociation in earlier time, the specific activity of PK was lower than the parent strain, as to HK, PFK, CS and Eno2p, the trend is quite another thing. Under the condition of oxidation and radiation, SOD and CAT activity of the mutant strain were down regulated, but the trehalose accumulation is up-regulated by the deletion of vacuolar proteinase A. It can be speculated that under the adversity, the absence of Saccharomyces cerevisiae proteinase A has a great regulation effect on the dynamic balance of trehalose synthesis and decomposition.