Sulfhydryl Oxidation, Not Disulfide Isomerization, Is the Principal Function of Protein Disulfide Isomerase in Yeast Saccharomyces cerevisiae*
Sulfhydryl Oxidation, Not Disulfide Isomerization, Is the Principal Function of Protein Disulfide Isomerase in Yeast Saccharomyces cerevisiae*
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DOI：
10.1074/jbc.m405640200
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发表时间：
2004-08
影响因子：
4.8
通讯作者：
A. Solovyov;Ruoyu Xiao;H. Gilbert
中科院分区：
文献类型：
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作者：
A. Solovyov;Ruoyu Xiao;H. Gilbert
蛋白质二硫键异构酶（PDI）是真核细胞内质网中一种重要的蛋白质折叠辅助酶，它催化蛋白质折叠过程中二硫键的形成（氧化酶活性）和可能不正确形成的二硫键的异构化（异构酶活性）。催化巯基-二硫键交换所需的PDI的细胞活力在酿酒酵母，但一直有一些不确定性，无论是在细胞中的PDI的基本作用是氧化酶或异构酶。我们研究了具有高氧化酶活性和非常低的异构酶活性的PDI构建体补充S.啤酒。酵母PDI的单个催化结构域（PDIa′）在体外具有野生型PDI的50%的氧化酶活性，但只有5%的异构酶活性。使用诱导型/阻遏型GAL 1 -10启动子滴定PDI的表达表明，维持正常生长速率所需的野生型PDI蛋白的量是从PDI 1染色体位置正常表达的量的60%或更多。单个催化结构域（PDIa′）所需的摩尔量大约是野生型PDI所需的摩尔量的两倍，野生型PDI含有两个催化结构域。这种比较表明，高（>60%）的PDI氧化酶活性对于酵母生长和活力是关键的，而需要少于6%的其异构酶活性。
Protein disulfide isomerase (PDI) is an essential protein folding assistant of the eukaryotic endoplasmic reticulum that catalyzes both the formation of disulfides during protein folding (oxidase activity) and the isomerization of disulfides that may form incorrectly (isomerase activity). Catalysis of thiol-disulfide exchange by PDI is required for cell viability in Saccharomyces cerevisiae, but there has been some uncertainty as to whether the essential role of PDI in the cell is oxidase or isomerase. We have studied the ability of PDI constructs with high oxidase activity and very low isomerase activity to complement the chromosomal deletion of PDI1 in S. cerevisiae. A single catalytic domain of yeast PDI (PDIa′) has 50% of the oxidase activity but only 5% of the isomerase activity of wild-type PDI in vitro. Titrating the expression of PDI using the inducible/repressible GAL1–10 promoter shows that the amount of wild-type PDI protein needed to sustain a normal growth rate is 60% or more of the amount normally expressed from the PDI1 chromosomal location. A single catalytic domain (PDIa′) is needed in molar amounts that are approximately twice as high as those required for wild-type PDI, which contains two catalytic domains. This comparison suggests that high (>60%) PDI oxidase activity is critical to yeast growth and viability, whereas less than 6% of its isomerase activity is needed.