Biosynthesis of the respiratory formate dehydrogenases from Escherichia coli: characterization of the FdhE protein
Biosynthesis of the respiratory formate dehydrogenases from Escherichia coli: characterization of the FdhE protein
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DOI：
10.1007/s00203-008-0420-4
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发表时间：
2008-12-01
影响因子：
2.8
通讯作者：
Sargent, Frank
中科院分区：
文献类型：
--
作者：
Lueke, Iris;Butland, Gareth;Sargent, Frank
大肠杆菌可以进行两种模式的甲酸盐代谢。在呼吸条件下，两种位于胞质周的甲酸脱氢酶同工酶将甲酸氧化与跨膜电化学梯度的产生偶联;在发酵条件下，第三种胞质同工酶参与甲酸向CO（2）和H（2）的转化。呼吸性甲酸脱氢酶是包含三个亚基的氧化还原酶：含钼辅因子和FeS簇的催化亚基;电子转移铁氧还蛋白;和膜整合细胞色素B。催化亚基及其铁氧还蛋白伴侣通过双精氨酸转运（达特）途径作为复合物靶向周质。这些酶的生物合成受称为FdhE的辅助蛋白的控制。本研究表明，E.大肠杆菌FdhE与呼吸性甲酸脱氢酶的催化亚基相互作用。重组FdhE的纯化表明，该蛋白是一种铁结合的红氧还蛋白，可以采用单体和同源二聚体形式。细菌双杂交分析表明FdhE的同型二聚体形式通过厌氧稳定。定点突变表明，保守的半胱氨酸基序是必不可少的FdhE蛋白的生理活性，也参与铁连接。
Escherichia coli can perform two modes of formate metabolism. Under respiratory conditions, two periplasmically-located formate dehydrogenase isoenzymes couple formate oxidation to the generation of a transmembrane electrochemical gradient; and under fermentative conditions a third cytoplasmic isoenzyme is involved in the disproportionation of formate to CO(2) and H(2). The respiratory formate dehydrogenases are redox enzymes that comprise three subunits: a molybdenum cofactor- and FeS cluster-containing catalytic subunit; an electron-transferring ferredoxin; and a membrane-integral cytochrome b. The catalytic subunit and its ferredoxin partner are targeted to the periplasm as a complex by the twin-arginine transport (Tat) pathway. Biosynthesis of these enzymes is under control of an accessory protein termed FdhE. In this study, it is shown that E. coli FdhE interacts with the catalytic subunits of the respiratory formate dehydrogenases. Purification of recombinant FdhE demonstrates the protein is an iron-binding rubredoxin that can adopt monomeric and homodimeric forms. Bacterial two-hybrid analysis suggests the homodimer form of FdhE is stabilized by anaerobiosis. Site-directed mutagenesis shows that conserved cysteine motifs are essential for the physiological activity of the FdhE protein and are also involved in iron ligation.