Structural Biology and NMR Laboratory, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200, Copenhagen N, Denmark.
Cell Biology and Physiology, Department of Biology, University of Copenhagen, Universitetsparken 13, DK-2100, Copenhagen Ø, Denmark.
Commun Biol. 2020 Dec 3;3(1):731. doi: 10.1038/s42003-020-01455-6.
Dynamic interactions of proteins with lipid membranes are essential regulatory events in biology, but remain rudimentarily understood and particularly overlooked in membrane proteins. The ubiquitously expressed membrane protein Na/H-exchanger 1 (NHE1) regulates intracellular pH (pH) with dysregulation linked to e.g. cancer and cardiovascular diseases. NHE1 has a long, regulatory cytosolic domain carrying a membrane-proximal region described as a lipid-interacting domain (LID), yet, the LID structure and underlying molecular mechanisms are unknown. Here we decompose these, combining structural and biophysical methods, molecular dynamics simulations, cellular biotinylation- and immunofluorescence analysis and exchanger activity assays. We find that the NHE1-LID is intrinsically disordered and, in presence of membrane mimetics, forms a helical αα-hairpin co-structure with the membrane, anchoring the regulatory domain vis-a-vis the transport domain. This co-structure is fundamental for NHE1 activity, as its disintegration reduced steady-state pH and the rate of pH recovery after acid loading. We propose that regulatory lipid-protein co-structures may play equally important roles in other membrane proteins.
蛋白质与脂质膜的动态相互作用是生物学中基本的调节事件，但在膜蛋白中仍然知之甚少，特别是被忽视了。普遍表达的膜蛋白 Na/H- exchanger 1（NHE1）通过调节细胞内 pH（pH）来发挥作用，其功能失调与癌症和心血管疾病等有关。NHE1 具有一个长的、调节性的胞质域，带有一个被描述为脂质相互作用域（LID）的近膜区域，然而，LID 的结构和潜在的分子机制尚不清楚。在这里，我们结合结构和生物物理方法、分子动力学模拟、细胞生物素化和免疫荧光分析以及交换活性测定来分解这些结构。我们发现，NHE1-LID 本质上是无序的，并且在存在膜类似物的情况下，与膜形成一个螺旋 αα-发夹共结构，相对于转运结构域锚定调节结构域。这种共结构对于 NHE1 的活性是至关重要的，因为它的解体降低了稳态 pH 值，并减少了酸加载后 pH 值恢复的速度。我们提出，调节性的脂质-蛋白质共结构可能在其他膜蛋白中同样起着重要的作用。
