Binding of 2'(3')-O-(2,4-6-trinitrophenyl) ADP to soluble alpha beta protomers of Na, K-ATPase modified with fluorescein isothiocyanate. Evidence for two distinct nucleotide sites.

Binding of 2'(3')-O-(2,4-6-trinitrophenyl) ADP to soluble alpha beta protomers of Na, K-ATPase modified with fluorescein isothiocyanate. Evidence for two distinct nucleotide sites.
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2(3)-O-(2,4-6-三硝基苯基) ADP 与异硫氰酸荧光素修饰的 Na,K-ATP 酶可溶性 αβ 原聚体的结合。

DOI:
10.1074/jbc.271.21.12317
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发表时间:
1996
期刊:
The Journal of biological chemistry
影响因子:
--
通讯作者:
J. Cavieres
J. Cavieres
中科院分区:
--
文献类型:
--
作者:
D. Ward;J. Cavieres

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Na,K-ATP 酶的明确溶解原体(一个 α 加一个 β 亚基)的总体反应保留了用膜结合酶观察到的双重 ATP 依赖性,在亚微摩尔和亚毫摩尔范围内具有独特的 ATP 效应(Ward, D. G. 和 Cavieres, J. D. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 5332-5336)。我们现在发现αβ原聚体的K+/-磷酸酶活性仍然被2'(3')-O-(2,4,6-三硝基苯基)腺苷5'-二磷酸(TNP-ADP)抑制。最重要的是,用异硫氰酸荧光素共价封闭高亲和力ATP位点的原体酶可以清楚地观察到TNP-ADP效应。我们得出结论,核苷酸可以结合在 Na,K-ATP 酶每个原体单元的两个离散位点上。
The overall reaction of well-defined solubilized protomers of Na,K-ATPase (one alpha plus one beta subunit) retains the dual ATP dependence observed with the membrane-bound enzyme, with distinctive ATP effects in the submicromolar and submillimolar ranges (Ward, D. G., and Cavieres, J. D. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 5332-5336). We have now found that the K+/-phosphatase activity of the alpha beta protomers is still inhibited by 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-diphosphate (TNP-ADP). What is most significant is that the TNP-ADP effect can be observed clearly with protomeric enzyme whose high affinity ATP site has been blocked covalently with fluorescein isothiocyanate. We conclude that nucleotides can bind at two discrete sites in each protomeric unit of Na,K-ATPase.
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