Neutral Phospholipids Stimulate Na,K-ATPase Activity A SPECIFIC LIPID-PROTEIN INTERACTION
Neutral Phospholipids Stimulate Na,K-ATPase Activity A SPECIFIC LIPID-PROTEIN INTERACTION
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DOI：
10.1074/jbc.m112.446997
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发表时间：
2013-04-05
影响因子：
4.8
通讯作者：
Karlish, Steven J. D.
中科院分区：
文献类型：
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作者：
Haviv, Haim;Habeck, Michael;Karlish, Steven J. D.
膜蛋白与磷脂的相互作用或者是通过跨膜片段周围的环状层，或者是通过特定的脂-蛋白相互作用。尽管在膜蛋白的许多晶体结构中观察到了特异结合的磷脂，但它们的作用尚不清楚。Na，K-ATPase高度依赖于酸性磷脂，尤其是磷脂酰丝氨酸，以往对纯化的洗涤剂可溶性重组Na，K-ATPase的研究表明，磷脂酰丝氨酸稳定并特异性地与蛋白质相互作用。最近，磷脂酰丝氨酸结合位点位于αTM8-10的跨膜片段和FXYD蛋白之间。本文报道了中性磷脂、磷脂酰胆碱或磷脂酰乙醇胺对纯化的人α1β1或α1β1FXYD1复合体的Na，K-ATPase活性的刺激作用。在磷脂酰丝氨酸存在的情况下，大豆磷脂酰胆碱将Na，K-ATPase的周转率从5483+/-144提高到7552+/-105(p<；0.0001)。用天然或人工合成的磷脂制备的α-1-β-1FXYD1络合物的分析表明，对含有多不饱和脂肪酰链的中性磷脂，特别是二异油酰磷脂酰胆碱或磷脂酰乙醇胺具有结构选择性的刺激作用。与磷脂酰丝氨酸相反，磷脂酰胆碱或磷脂酰乙醇胺会破坏Na，K-ATPase的稳定性。中性磷脂对Na，K-ATPase活性的刺激和不稳定的结构选择性将这些作用与磷脂酰丝氨酸的稳定作用区分开来，暗示磷脂结合在不同的位置。对鲨鱼Na，K-ATPase电子密度的重新检测与位于跨膜片段αTM8-10和TMFXYD(A位点)和TM2-4，-6-和9(B位点)之间的两个结合磷脂是一致的。比较E_2和E_1构象中的磷脂结合口袋，提示中性磷脂刺激Na，K-ATPase活性的可能机制。
Membrane proteins interact with phospholipids either via an annular layer surrounding the transmembrane segments or by specific lipid-protein interactions. Although specifically bound phospholipids are observed in many crystal structures of membrane proteins, their roles are not well understood. Na,K-ATPase is highly dependent on acid phospholipids, especially phosphatidylserine, and previous work on purified detergent-soluble recombinant Na, K-ATPase showed that phosphatidylserine stabilizes and specifically interacts with the protein. Most recently the phosphatidylserine binding site has been located between transmembrane segments of alpha TM8-10 and the FXYD protein. This paper describes stimulation of Na, K-ATPase activity of the purified human alpha 1 beta 1 or alpha 1 beta 1FXYD1 complexes by neutral phospholipids, phosphatidylcholine, or phosphatidylethanolamine. In the presence of phosphatidylserine, soy phosphatidylcholine increases the Na, K-ATPase turnover rate from 5483 +/- 144 to 7552 +/- 105 (p < 0.0001). Analysis of alpha 1 beta 1FXYD1 complexes prepared with native or synthetic phospholipids shows that the stimulatory effect is structurally selective for neutral phospholipids with polyunsaturated fatty acyl chains, especially dilinoleoyl phosphatidylcholine or phosphatidylethanolamine. By contrast to phosphatidylserine, phosphatidylcholine or phosphatidylethanolamine destabilizes the Na, K-ATPase. Structural selectivity for stimulation of Na, K-ATPase activity and destabilization by neutral phospholipids distinguish these effects from the stabilizing effects of phosphatidylserine and imply that the phospholipids bind at distinct sites. A re-examination of electron densities of shark Na, K-ATPase is consistent with two bound phospholipids located between transmembrane segments alpha TM8-10 and TMFXYD(site A) and between TM2, -4, -6, -and 9 (site B). Comparison of the phospholipid binding pockets in E2 and E1 conformations suggests a possible mechanism of stimulation of Na, K-ATPase activity by the neutral phospholipid.