Emmermann M, Clericus M, Braun H P, Mozo T, Heins L, Kruft V, Schmitz U K
Institut für Genbiologische Forschung Berlin GmbH, Germany.
Plant Mol Biol. 1994 May;25(2):271-81. doi: 10.1007/BF00023243.
The mitochondrial iron-sulfur protein (also termed Rieske iron-sulfur protein) of cytochrome c reductase was purified from potato tubers and identified with heterologous antibodies. The sequences of the N-terminus of this 25 kDa protein and of an internal peptide were determined to design oligonucleotide mixtures for screening a cDNA library. One class of cDNA clones containing an open reading frame of 265 amino acids was isolated. The encoded protein contains the peptide sequences of the 25 kDa protein and shares about 50% sequence identity with the Rieske iron-sulfur proteins from fungi and around 43% with those from mammals. In vitro transcription and translation of the cDNA reveals that the iron-sulfur protein is made as a larger precursor of 30 kDa which is processed by the cytochrome c reductase/processing peptidase complex from potato. The processing product obtained after in vitro processing has the same size as the mature protein imported into isolated mitochondria. The presequence, which targets the protein to the organelle, is 53 amino acids long and has molecular features different from those found in presequences of fungal iron-sulfur proteins, which are processed in two steps. Our results indicate that, unlike in yeast and Neurospora, the presequence of the iron-sulfur protein from potato is removed by a single processing enzyme in one step.
细胞色素c还原酶的线粒体铁硫蛋白（也称为里斯克铁硫蛋白）从马铃薯块茎中纯化出来，并用异源抗体进行鉴定。测定了这种25 kDa蛋白N端和一个内部肽段的序列，以设计用于筛选cDNA文库的寡核苷酸混合物。分离出一类包含265个氨基酸开放阅读框的cDNA克隆。编码的蛋白包含25 kDa蛋白的肽段序列，与真菌的里斯克铁硫蛋白有大约50%的序列同一性，与哺乳动物的该蛋白有大约43%的序列同一性。cDNA的体外转录和翻译表明，铁硫蛋白最初是一个30 kDa的较大前体，由马铃薯的细胞色素c还原酶/加工肽酶复合体进行加工。体外加工后得到的加工产物与导入分离线粒体的成熟蛋白大小相同。将该蛋白靶向细胞器的前导序列长53个氨基酸，其分子特征与真菌铁硫蛋白前导序列不同，后者是分两步加工的。我们的结果表明，与酵母和粗糙脉孢菌不同，马铃薯铁硫蛋白的前导序列由一种单一的加工酶一步去除。
