Binding of cardiotonic steroids to Na+,K+-ATPase in the E2P state
Binding of cardiotonic steroids to Na+,K+-ATPase in the E2P state
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DOI：
10.1073/pnas.2020438118
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发表时间：
2021-01-05
影响因子：
11.1
通讯作者：
Toyoshima, Chikashi
中科院分区：
文献类型：
--
作者：
Kanai, Ryuta;Cornelius, Flemming;Toyoshima, Chikashi
钠泵（Na+，K+-ATP酶，NKA）对动物细胞至关重要，因为它积极维持Na+和K+跨细胞膜的电化学梯度。它是强心类固醇（CTS）如哇巴因和地高辛的靶点。由于CTS几乎是唯一的NKA特异性强抑制剂，因此已开发了广泛的衍生物用于潜在的治疗用途。NKA-CTS复合物的几种晶体结构已被公布，但它们未能解释各种CTS的很大不同的抑制特性。例如，尽管CTS被认为通过结合E2 P状态的NKA来抑制ATP酶活性，但我们不知道结合是否伴随有大的构象变化，因为没有晶体结构可用于不含CTS的E2 P状态。在这里，我们描述的BeF 3-复合物的NKA代表E2 P基态的晶体结构，然后8个晶体结构的7个CTS，包括rostafuroxin和istaroxime，两个新的成员在临床试验中，在复杂的NKA在E2 P状态。NKA的构象几乎是相同的，在所有的复合物与和没有CTS，表明CTS绑定到一个预先形成的空腔在NKA。通过比较在四种不同条件下测量的CTS的抑制效力，我们阐明了CTS的不同结构特征如何导致不同的抑制特性。晶体结构也解释了K+（+）-拮抗作用，并提出了一种途径亚型特异性CTS。
The sodium pump (Na+, K+-ATPase, NKA) is vital for animal cells, as it actively maintains Na+ and K+ electrochemical gradients across the cell membrane. It is a target of cardiotonic steroids (CTSs) such as ouabain and digoxin. As CTSs are almost unique strong inhibitors specific to NKA, a wide range of derivatives has been developed for potential therapeutic use. Several crystal structures have been published for NKA-CTS complexes, but they fail to explain the largely different inhibitory properties of the various CTSs. For instance, although CTSs are thought to inhibit ATPase activity by binding to NKA in the E2P state, we do not know if large conformational changes accompany binding, as no crystal structure is available for the E2P state free of CTS. Here, we describe crystal structures of the BeF3- complex of NKA representing the E2P ground state and then eight crystal structures of seven CTSs, including rostafuroxin and istaroxime, two new members under clinical trials, in complex with NKA in the E2P state. The conformations of NKA are virtually identical in all complexes with and without CTSs, showing that CTSs bind to a preformed cavity in NKA. By comparing the inhibitory potency of the CTSs measured under four different conditions, we elucidate how different structural features of the CTSs result in different inhibitory properties. The crystal structures also explain K+(+)-antagonism and suggest a route to isoform specific CTSs.