氧化胁迫对酿酒酵母NAD(H)激酶突变体呼吸链活性的影响

摘要

[目的]了解酿酒酵母线粒体NAD(H)激酶Pos5p对呼吸链活性的维持是否与其抗氧化功能有关.[方法]比较在不同类型的氧化胁迫试剂作用下,野生菌BY4742、POS5基因缺失体pos5△及其回补体pos5△/POS5-YEp的呼吸链各个酶复合体的活性变化及细胞内活性氧水平变化.[结果]在非胁迫条件下,pos5△的各个复合体活性明显低于BY4742,而pos5△/POS5-YEp的活性有所恢复,这与它们的胞内活性氧水平相一致.在甲萘醌胁迫下,BY4742和pos5△的各个复合体活性都发生不同程度的下降,但pos5△/POS5-YEp的活性都升高.在H2O2、马来酸二乙酯胁迫下,除个别复合体外,BY4742、pos5△和pos5△/POS5-YEp的呼吸链复合体活性都降低,尤以pos5△的活性降低最为严重,BY4742的活性降低则较少,而pos5△/POS5-YEp在H2O2胁迫下的活性降低得到了缓解.说明甲萘醌、H2O2和马来酸二乙酯胁迫会造成酿酒酵母呼吸链各个复合体发生损伤,而过表达Pos5p则有助于缓解甲萘醌和H2O2引起的损伤.[结论]Pos5p对呼吸链的作用与其抗氧化功能有相关性.%[Objective] To realize the relationship between respiratory maintenance and anti-oxidative function of Pos5p, the mitochondrial NAD(H) kinase in Saccharomyces cerevisiae.[Methods] The respiratory chain activity and reactive oxygen species (ROS) level of wild-type BY4742, POS5 gene deletion mutant pos5△, and pos5△ containing POS5-YEplac195 plasmid (pos5△/POS5-YEp) under exposure to different kinds of oxidative reagents were compared.[Results] At the normal growth condition, pos5△ exhibited poorer respiratory chain activity than that of BY4742, while pos5△/POS5-YEp gave the partial restored activity, consistent with the ROS level of these three strains.Under the oxidative stress of menadione, the respiratory chain activities of BY4742 and pos5△ decreased, while that of pos5△/POS5-YEp increased.Under the oxidative stress of H2O2 and diethyl maleate, nearly all of the activities of BY4742, pos5△ and pos5△/POS5-YEp reduced, with that of pos5△ most seriously, and that of pos5△/POS5-YEp under H2O2 lessened.Thus indicated that the oxidative stress would injury the respiratory chain complex of S.cerevisiae and over-expression of Pos5p could alleviate the injury from menadione and H2O2.[Conclusion] The protecting function of Pos5p on respiratory chain was correlated to its antioxidative defense function.