Assignment of the CO-sensitive carboxyl group in mitochondrial forms of cytochrome c oxidase using yeast mutants
Assignment of the CO-sensitive carboxyl group in mitochondrial forms of cytochrome c oxidase using yeast mutants
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DOI：
10.1016/j.bbabio.2012.03.036
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发表时间：
2012-10-01
影响因子：
4.3
通讯作者：
Rich, Peter R.
中科院分区：
文献类型：
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作者：
Marechal, Amandine;Meunier, Brigitte;Rich, Peter R.
将E243D和I67N的点突变引入酿酒酵母线粒体细胞色素C氧化酶的Iota 6组氨酸标记亚单位(6H-WT)。对两个突变体(6H-E243D(I)和6H-I67N(I))进行了纯化，分别相当于6H-WT周转率的50%和10%。6H-WT的光致CO光解FTIR差谱在1749/1740 cm(-1)处出现峰/谷，与牛CcO一样，D2O中峰/谷下移了7 cm(-1)。在6H-E243D(I)中，条带移动到1736/1762 cM(-1)，证实线粒体CCOs中受CO结合影响的羧基是E243。在6H-I67N(I)中，1740 cm(-1)处的谷移至1743 cm(-1)，伴随的峰强度大大降低。这证实了I67N突变干扰了E243周围的构象变化。本文是题为：第17届欧洲生物能量学会议(EBEC 2012)特刊的一部分。(C)2012爱思唯尔B.V.保留所有权利。
Point mutations of E243D and I67N were introduced into subunit Iota 6histidine-tagged (6H-WT) form of yeast Saccharomyces cerevisiae mitochondrial cytochrome c oxidase. The two mutants (6H-E243D(I) and 6H-I67N(I)) were purified and showed approximate to 50 and 10% of the 6H-WT turnover number. Light-induced CO photolysis FTIR difference spectra of the 6H-WT showed a peak/trough at 1749/1740 cm(-1), as seen in bovine CcO, which downshifted by 7 cm(-1) in D2O. The bands shifted to 1736/1762 cm(-1) in 6H-E243D(I), establishing that the carboxyl group affected by CO binding in mitochondrial CcOs is E243. In 6H-I67N(I), the trough at 1740 cm(-1) was shifted to 1743 cm(-1) and its accompanying peak intensity was greatly reduced. This confirms that the I67N mutation interferes with conformational alterations around E243. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012). (C) 2012 Elsevier B.V. All rights reserved.