The Role Of Saccharomyces Cerevisiae Nadh Kinase Pos5p On The Defense Of Oxidative Stress

NADPH is one of the important coenzymes in cells, not only for the supply of reducing equivalents in biosynthetic pathways, but also for the protection of cells against the toxicity of ROS. In Saccharomyces cerevisiae, a mitochondrial NADH kinase encoded by POS5 gene is the major source of mitochondrial NADPH. Pos5p plays an important role on the maintenance of normal mitochondrial functions. Absence of POS5 gene leads to antioxidative dysfunction and respiratory defects of cells. However, the reason for the antioxidative dysfunction and respiration deficiency of pos5Δis still to be realized.In order to realize the relation of Pos5p with anti-oxidative system of yeast cells, we compared the growth phenotypes and intracellular reactive oxygen species (ROS) concentration of pos5Δwith some anti-oxidative gene deletion mutants. The results showed that pos5Δhad obvious growth defect under all of the assayed oxidative stress, that was menadione (VK3), hydrogen peroxide (H2O2) and diethyl maleate (DEM), whereas the anti-oxidative gene deletion mutants only had growth defect under their corresponding oxidative stress. Deletion of POS5 leads to the significantly higher level of intracellular ROS, while deletion of double anti-oxidative genes, that were sod2glr1Δand trr2glr1Δ, did not lead to so higher level of intracellular ROS. Under exposure to VK3, H2O2 and DEM, Pos5p could also help to scavenge intracellular ROS in varying degrees. This proved the importance of Pos5p on the defense of several kinds of oxidative stress and on the removal of intracellular ROS.Then we determined the intracellular coenzyme concentration of pos5Δ, that was NAD+, NADH, NADP+ and NADPH concentration. The result confirmed that Pos5p had a crucial role on the supply of intracellular NADPH. While under the exposure to VK3, H2O2 and DEM, Pos5p could catalyze the convertion of NAD(H) to NADP(H) by its NADH kinase activity and supply the NADP(H) exhausted, which indicated that Pos5p could protect cells from the injury of different kinds of oxidative reagents by its NADH kinase activity.Finally, we determined the respiratory chain complexes activities of pos5Δunder exposure to different kinds of oxidative stress, including complexⅡ, complexⅢ, complexⅡ+Ⅲand complexⅣactivities. The result indicated that the enzyme activities of complexⅡand complexⅣof POS5 gene deletion mutant were significantly lower than that of the wild-type strain. Under exporsure to H2O2 and DEM, most activities of respiratory chain complexes of pos5Δdecreased, while under exporsure to VK3, the activities sharply increased. The possible adaptive response mechanism remains to be further researched.S. cerevisiae is not only an important industrial microorganism, but also a typical eukaryotic model organism. In this paper, we studied the function of NADH kinase Pos5p on resistance to oxidative stress and confirmed that Pos5p has an important role on anti-oxidant system and on resistance to different kinds of oxidative reagents. Such study would be useful for us to comprehensively learn about the antioxidative mechanism of S. cerevisiae, and help us to improve antioxidative property of yeast cells.