Phillips J D, Graham L A, Trumpower B L
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755.
J Biol Chem. 1993 Jun 5;268(16):11727-36.
Deletion of QCR9, the nuclear gene encoding the 7.3-kDa subunit 9 of the cytochrome bc1 complex, impairs respiration of Saccharomyces cerevisiae, coincident with loss of ubiquinol-cytochrome c oxidoreductase activity. Optical spectra of mitochondrial membranes from yeast in which the gene for subunit 9 is deleted show a diminution of cytochrome b absorption similar to the spectra of membranes from yeast in which the gene for the Rieske iron-sulfur protein is deleted, suggesting an interaction between subunit 9, iron-sulfur protein, and cytochrome b. Synthesis of cytochrome b by mitochondria from the deletion strain is unimpaired, indicating that the diminished b absorption is due to a post-assembly effect on the heme environment resulting from the absence of subunit 9. Iron-sulfur protein is present in normal amounts and processed to its mature form in the absence of subunit 9, although the protein is more labile to endogenous proteases during the isolation of membranes. EPR spectroscopy of membranes from the subunit 9 deletion strain indicates that the g = 1.90 signal characteristic of the Rieske iron-sulfur cluster is absent, even though mature sized apoprotein is present. Pre-steady state reduction of cytochrome c1 is markedly slowed, but not eliminated, in the subunit 9 deletion strain, which suggests that an EPR-silent, sluggishly reactive derivative of the iron-sulfur cluster is present. These results suggest that in the absence of subunit 9 the conformation of iron-sulfur protein is altered such that the protein is more labile, the iron-sulfur cluster is not properly inserted, and iron-sulfur protein interaction with cytochrome b is modified in a manner which distorts the heme environment. This is the first instance in which deletion of one of the supernumerary subunits of the cytochrome bc1 complex results in the loss of function of a redox center within the complex, without a concomitant loss of other subunits.
细胞色素bc1复合体7.3 kDa亚基9的编码核基因QCR9的缺失，会损害酿酒酵母的呼吸作用，这与泛醇 - 细胞色素c氧化还原酶活性的丧失同时发生。在9号亚基基因被删除的酵母线粒体膜的光谱中，细胞色素b的吸收减少，这与铁硫蛋白基因被删除的酵母膜的光谱相似，表明9号亚基、铁硫蛋白和细胞色素b之间存在相互作用。缺失菌株的线粒体合成细胞色素b不受影响，这表明b吸收减少是由于9号亚基缺失导致的血红素环境的组装后效应。尽管在膜分离过程中铁硫蛋白对内源蛋白酶更不稳定，但在没有9号亚基的情况下，铁硫蛋白的含量正常并加工成其成熟形式。9号亚基缺失菌株的膜的电子顺磁共振光谱表明，即使存在成熟大小的脱辅基蛋白，里氏铁硫簇特有的g = 1.90信号也不存在。在9号亚基缺失菌株中，细胞色素c1的稳态前还原明显减慢，但并未消除，这表明存在一种电子顺磁共振沉默、反应迟缓的铁硫簇衍生物。这些结果表明，在没有9号亚基的情况下，铁硫蛋白的构象发生改变，使得该蛋白更不稳定，铁硫簇不能正确插入，并且铁硫蛋白与细胞色素b的相互作用被改变，从而扭曲了血红素环境。这是细胞色素bc1复合体多余亚基之一的缺失导致复合体内氧化还原中心功能丧失，而其他亚基没有同时丧失的首个实例。
