His(15) of subunit a of the Escherichia coli ATP synthase is important for the structure or assembly of the membrane sector F(o).
His(15) of subunit a of the Escherichia coli ATP synthase is important for the structure or assembly of the membrane sector F(o).
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大肠杆菌 ATP 合酶 a 亚基的 His(15) 对于膜扇区 F(o) 的结构或组装很重要。
DOI：
10.1006/abbi.1999.1306
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发表时间：
1999
影响因子：
3.9
通讯作者：
Vik,SB
中科院分区：
文献类型：
--
作者：
Patterson,AR;Wada,T;Vik,SB
大肠杆菌ATP合酶a亚基氨基端约37个氨基酸定位于周质。结果表明，一个单一的氨基酸取代，H15 D，破坏亚基的组装，并导致ATP合酶功能的丧失。在这项研究中，9个氨基酸的保守区域，11-19，最初随机诱变，产生没有突变体，可以在琥珀酸盐基本培养基上生长。随后的诱变，局限于残基His 14，His 15和Asn 17，表明含有H15 D的构建体是最有害的。构建并分析了四种单突变体：H15 A、H14 D、H15 A和H15 D。只有H15 D显着受损，相对于ATP驱动的质子易位，被动质子渗透性通过Fo，和膜结合ATP酶的敏感性DCCD。免疫印迹分析表明H15 D的亚基a水平非常低。在位置14、15、17和18处构建半胱氨酸突变。残基14、15和17显示在周质空间中可接近，而残基18不可接近，表明该区域稳定折叠。虽然His 14和His 15在一组细菌中是保守的，但这里的结果表明它们是不等价的，并且支持His 15在ATP合酶的组装或结构中的特定作用。
Approximately 37 amino acids at the amino-terminus of subunit a of the Escherichia coli ATP synthase are found localized to the periplasm. Results indicate that a single amino acid substitution, H15D, disrupts assembly of subunit a and causes a loss of ATP synthase function. In this study, a conserved region of nine amino acids, 11–19, was initially mutagenized randomly, generating no mutants that could grow on succinate-minimal medium. Subsequent mutagenesis, confined to residues His14, His15, and Asn17, indicated that constructs containing H15D were the most deleterious. Four single mutants were constructed and analyzed: H15A, H14D, H15A, and H15D. Only H15D was significantly impaired, with respect to ATP-driven proton translocation, passive proton permeability through Fo, and sensitivity of membrane-bound ATPase to DCCD. Immunoblot analysis indicated very low levels of subunit a from H15D. Cysteine mutations were constructed at positions 14, 15, 17, and 18. Residues 14, 15, and 17 were shown to be accessible in the periplasmic space, while residue 18 was not, indicating that this region was stably folded. While both His14and His15are conserved among a group of bacteria, results presented here indicate that they are not equivalent, and that a specific role for His15in the assembly or structure of the ATP synthase is supported.