Braun H P, Emmermann M, Kruft V, Bödicker M, Schmitz U K
Institut für Genbiologische Forschung Berlin GmbH, Germany.
Planta. 1995;195(3):396-402. doi: 10.1007/BF00202597.
The bc1-complex (EC 1.10.2.2.) from Triticum aestivum L. was purified by cytochrome-c affinity chromatography and gel filtration using either etiolated seedlings or wheat-germ extract as starting material. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the isolated enzyme revealed ten bands, which were analysed by immunoblotting and direct amino-acid sequencing. The enzyme from wheat is the first bc1-complex that is reported to contain four core proteins (55.5, 55.0, 51.5 and 51.0 kDa). In addition, the wheat bc1-complex comprises cytochrome b (35 kDa), cytochrome c1 (33 kDa) the "Rieske" iron-sulphur protein (25 kDa) and three small subunits < 15 kDa. This composition differs from the one reported in fungi, mammals and potato. Partial sequence determination of the large subunits suggests that the 55.5- and 55.0-kDa-proteins represent the beta-subunit of the general mitochondrial processing peptidase, and the 51.5- and 51.0-kDa proteins the alpha-subunit of this enzyme. The bc1-complex from wheat efficiently processes mitochondrial precursor proteins as shown in an in-vitro processing assay. In control experiments the isolated bc1-complexes from potato, yeast, Neurospora and beef, all purified by the same isolation procedure, were also tested for processing activity. Only the protein complexes from plants contain the general mitochondrial processing peptidase. The composition of the wheat bc1-complex sheds new light on the co-evolution of the processing peptidase and the middle segment of the respiratory chain.
以黄化幼苗或小麦胚芽提取物为起始材料，通过细胞色素c亲和层析和凝胶过滤法对普通小麦（Triticum aestivum L.）的bc1复合体（EC 1.10.2.2.）进行了纯化。对分离得到的酶进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳，显示出十条带，通过免疫印迹和直接氨基酸测序对其进行了分析。小麦中的这种酶是首个被报道含有四种核心蛋白（55.5、55.0、51.5和51.0 kDa）的bc1复合体。此外，小麦bc1复合体包含细胞色素b（35 kDa）、细胞色素c1（33 kDa）、“里氏”铁硫蛋白（25 kDa）和三个小于15 kDa的小亚基。这种组成与真菌、哺乳动物和马铃薯中报道的不同。大亚基的部分序列测定表明，55.5 kDa和55.0 kDa的蛋白代表线粒体加工肽酶的β亚基，而51.5 kDa和51.0 kDa的蛋白代表该酶的α亚基。如体外加工试验所示，小麦的bc1复合体能够有效地加工线粒体前体蛋白。在对照实验中，还对通过相同分离程序纯化的来自马铃薯、酵母、粗糙脉孢菌和牛肉的分离bc1复合体的加工活性进行了测试。只有植物来源的蛋白复合体含有线粒体加工肽酶。小麦bc1复合体的组成揭示了加工肽酶与呼吸链中间段共同进化的新线索。
