Isolation and characterization of a cDNA clone for bovine cytochrome c oxidase subunit IV.
Isolation and characterization of a cDNA clone for bovine cytochrome c oxidase subunit IV.
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牛细胞色素 c 氧化酶亚基 IV cDNA 克隆的分离和表征。
DOI：
10.1073/pnas.81.20.6295
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发表时间：
1984
影响因子：
11.1
通讯作者：
Grossman,LI
中科院分区：
文献类型：
--
作者：
Lomax,MI;Bachman,NJ;Nasoff,MS;Caruthers,MH;Grossman,LI
我们分离了牛细胞色素 c 氧化酶（铁细胞色素 c：氧氧化还原酶，EC 1.9.3.1）亚基 IV 前体的 cDNA 克隆。通过将 cDNA 插入质粒载体 pBR322，从成年牛肝的 Poly(A)+ RNA 构建 cDNA 文库。通过与[32P]标记的合成寡脱氧核糖核苷酸的两种混合物进行菌落杂交来筛选转化体。我们筛选了20,000个具有与编码氨基酸98-103的所有16种可能序列互补的七聚体混合物的转化体，并获得了编码亚基IV氨基酸序列的两个cDNA克隆。我们确定了较大插入片段（416 个碱基对）的 DNA 序列，其中包含成熟蛋白质的氨基酸 1-107 的编码序列和 NH2 末端延伸（前序列）。成熟蛋白的推导氨基酸序列与之前确定的蛋白序列相同：NH2 末端延伸序列在加工蛋白的 NH2 末端的氨基酸 -22 处含有潜在的引发剂蛋氨酸。前序列非常基本，包含多种精氨酸，其中一种位于加工位点。不存在与细菌和真核信号肽中发现的类似的疏水区域，但与其他线粒体蛋白前序列有同源性，这可能包括其目的地和处理的共同信号。
We have isolated a cDNA clone for the precursor to subunit IV of bovine cytochrome c oxidase (ferrocytochrome c:oxygen oxidoreductase, EC 1.9.3.1). A cDNA library was constructed from poly(A)+ RNA of adult beef liver by insertion of cDNA into the plasmid vector pBR322. Transformants were screened by colony hybridization with two mixtures of [32P]-labeled synthetic oligodeoxyribonucleotides. We screened 20,000 transformants with a mixture of heptadecamers complementary to all 16 possible sequences encoding amino acids 98-103 and obtained two cDNA clones encoding subunit IV amino acid sequences. We determined the DNA sequence of the larger (416 base-pair) insert, which contains the coding sequence for amino acids 1-107 of the mature protein and an NH2-terminal extension (presequence). The deduced amino acid sequence of the mature protein is identical with the previously determined protein sequence: the sequence of the NH2-terminal extension contains a potential initiator methionine at amino acid -22 from the NH2-terminus of the processed protein. The presequence is quite basic and contains several arginines, including one at the processing site. No hydrophobic region analogous to that found in bacterial and eukaryotic signal peptides is present, but there are homologies with other mitochondrial protein presequences, which may include a common signal for their destination and processing.
基因片段的化学和酶法合成
DOI：
--
发表时间：
1983
期刊：
影响因子：
--
作者：
B. Roques
通讯作者：
B. Roques