Domains near ATP gamma phosphate in the catalytic site of H+-ATPase. Model proposed from mutagenesis and inhibitor studies.
Domains near ATP gamma phosphate in the catalytic site of H+-ATPase. Model proposed from mutagenesis and inhibitor studies.
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H-ATP 酶催化位点中 ATP γ 磷酸附近的结构域。
DOI：
10.1016/s0021-9258(18)53672-3
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
M. Futai
中科院分区：
文献类型：
--
作者：
A. Iwamoto;M. Park;M. Maeda;M. Futai
β - Gly-149残基位于大肠杆菌H(+)-ATP合成酶β亚基的一个富含甘氨酸的序列（Gly-Gly-Ala-Gly-Val-Gly-Lys-Thr；残基149-156）。用Ser替代β - Gly-149抑制β - Ser-174 - >Phe突变的作用(Iwamoto， A., Omote， H., Hanada， H., Tomioka， N., Itai， A., Maeda， M., and Futai， M. (1991) J. Biol。化学，266,16350-16355)，表明β Gly-149位于β Ser-174附近。在这项研究中，我们在149号位置引入了不同的残基，发现单个突变体β Cys-149存在缺陷。β - Gly-172 - >Glu、β - Ser-174 - >Phe、β - Glu-192 - >Val或β - Val-198 - >Ala替代均可抑制β - Cys-149突变的影响。这些结果表明β - Gly-149、β - Gly-172、β - Ser-174、β - Glu-192和β - Val-198残基在催化位点上的位置很近。根据这些发现，我们提出了ATP γ磷酸部分附近酶的催化位点的模型。具有双突变β - Cys-149/ β - Glu-172、β - Cys-149/ β - ph -174、β - Cys-149/ β - Val-192和β - Cys-149/ β - Ala-198的F1酶对双环己基碳二亚胺和腺苷三磷酸吡啶醇（ATP与β - Lys-155或β - Lys-201的ε -氨基形成希夫碱的亲和类似物）的敏感性低于野生型F1，并以ATP保护的方式对n-乙基马来酰亚胺敏感。这些抑制剂研究的结果与所提出的模型一致。
The beta Gly-149 residue is in a glycine-rich sequence (Gly-Gly-Ala-Gly-Val-Gly-Lys-Thr; residues 149-156) of the Escherichia coli H(+)-ATPase (ATP synthase) beta subunit. Substitution of beta Gly-149 by Ser suppressed the effect of the beta Ser-174–>Phe mutation (Iwamoto, A., Omote, H., Hanada, H., Tomioka, N., Itai, A., Maeda, M., and Futai, M. (1991) J. Biol. Chem. 266, 16350-16355), suggesting that beta Gly-149 is located near beta Ser-174. In this study, we introduced different residues at position 149 and found that a single mutant beta Cys-149 was defective. The effect of beta Cys-149 mutation was suppressed by beta Gly-172–>Glu, beta Ser-174–>Phe, beta Glu-192–>Val, or beta Val-198–>Ala replacement. These results suggest that beta Gly-149, beta Gly-172, beta Ser-174, beta Glu-192, and beta Val-198 residues are located close together in the catalytic site. From these findings we propose a model of the catalytic site of the enzyme near the gamma phosphate moiety of ATP. F1 enzymes with the double mutations beta Cys-149/beta Glu-172, beta Cys-149/beta Phe-174, beta Cys-149/beta Val-192, and beta Cys-149/beta Ala-198 were less sensitive than wild-type F1 to dicyclohexylcarbodiimide and adenosine triphosphopyridoxal (an affinity analogue of ATP forming a Schiff base with the epsilon-amino group of beta Lys-155 or beta Lys-201), and became sensitive to N-ethylmaleimide in an ATP-protected manner. These results of inhibitor studies are consistent with the proposed model.