Mutations Phe785Leu and Thr618Met in Na+, K+-ATPase, associated with familial rapid-onset dystonia parkinsonism, interfere with Na+ interaction by distinct mechanisms
Mutations Phe785Leu and Thr618Met in Na+, K+-ATPase, associated with familial rapid-onset dystonia parkinsonism, interfere with Na+ interaction by distinct mechanisms
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DOI：
10.1074/jbc.m601780200
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发表时间：
2006-07-07
影响因子：
4.8
通讯作者：
Vilsen, Bente
中科院分区：
文献类型：
--
作者：
Rodacker, Vivien;Toustrup-Jensen, Mads;Vilsen, Bente
Na+，K+-ATPase在脑功能中起着关键作用。最近发现Na+，K+-ATPase错义突变与家族性速发型肌张力障碍帕金森综合征(FRDP)相关。在这里，我们已经表征了FRDP突变Phe785Leu和Thr618Met的功能后果。这两个突变都会导致功能改变但活性的Na+，K+泵，表现出对细胞质Na+的表观亲和力降低，但潜在的机制在突变体之间有所不同。在Phe785Leu中，E-1与Na+的相互作用是缺陷的，E-1-E-2平衡没有移位。在Thr618Met中，由于构象平衡向K+封闭的E-2(K-2)形式移动，Na+亲和力降低。在两个突变体中，E2P磷酸酶外部激活部位的K+相互作用是正常的。在携带Phe785Leu或Thr618Met突变的患者中，细胞Na+稳态的改变可能是导致FRDP发生的主要因素。此外，Phe785Leu还干扰了胞外Na+的相互作用，并显著降低了哇巴因的亲和力。对另外两个Phe(785)突变体Phe785Leu/Leu786Phe和Phe785Tyr的分析表明，侧链的芳香功能及其确切位置对Na+和哇巴因的结合至关重要。结构模拟可以解释取代Phe(785)的影响，表明Phe(785)参与了三个跨膜片段之间的疏水网络。Thr(618)位于催化中心附近的细胞质部分，结构模拟表明Thr618Met突变干扰了E1型催化中心的键模式，从而使E-1相对于E-2(K-2)不稳定。
The Na+, K+-ATPase plays key roles in brain function. Recently, missense mutations in the Na+, K+-ATPase were found associated with familial rapid-onset dystonia parkinsonism (FRDP). Here, we have characterized the functional consequences of FRDP mutations Phe785Leu and Thr618Met. Both mutations lead to functionally altered, but active, Na+, K+-pumps, that display reduced apparent affinity for cytoplasmic Na+, but the underlying mechanism differs between the mutants. In Phe785Leu, the interaction of the E-1 form with Na+ is defective, and the E-1-E-2 equilibrium is not displaced. In Thr618Met, the Na+ affinity is reduced because of displacement of the conformational equilibrium in favor of the K+-occluded E-2(K-2) form. In both mutants, K+ interaction at the external activating sites of the E2P phosphoenzyme is normal. The change of cellular Na+ homeostasis is likely a major factor contributing to the development of FRDP in patients carrying the Phe785Leu or Thr618Met mutation. Phe785Leu moreover interferes with Na+ interaction on the extracellular side and reduces the affinity for ouabain significantly. Analysis of two additional Phe(785) mutants, Phe785Leu/Leu786Phe and Phe785Tyr, demonstrated that the aromatic function of the side chain, as well as its exact position, is critical for Na+ and ouabain binding. The effects of substituting Phe(785) could be explained by structural modeling, demonstrating that Phe(785) participates in a hydrophobic network between three transmembrane segments. Thr(618) is located in the cytoplasmic part of the molecule near the catalytic site, and the structural modeling indicates that the Thr618Met mutation interferes with the bonding pattern in the catalytic site in the E1 form, thereby destabilizing E-1 relative to E-2(K-2).