Jordan C, Püschel B, Koob R, Drenckhahn D
Institute of Anatomy, University of Würzburg, Germany.
J Biol Chem. 1995 Dec 15;270(50):29971-5. doi: 10.1074/jbc.270.50.29971.
Cytoskeleton membrane associations are important for a variety of cellular functions. The anion exchanger of erythrocytes (AE1) and Na+,K(+)-ATPase of polarized epithelial cells provide well studied examples of how integral membrane proteins are anchored via the linker molecule ankyrin to the spectrin-based membrane cytoskeleton. In the present study we have generated several recombinant fragments of the large (third) cytoplasmic domain (CD3) of Na+,K(+)-ATPase to define binding sites of ankyrin on CD3 at a molecular level. We provide evidence that a cluster of four amino acids, ALLK, is essential for binding of ankyrin to both recombinant CD3 and to native Na+,K(+)-ATPase. Once bound, conformational changes might uncover further binding sites for ankyrin on Na+,K(+)-ATPase. A motif related to the ALLK cluster is also present in the cytoplasmic domain of AE1 where this sequence (ALLLK) turned out to be also important for ankyrin binding. These motifs are highly conserved during evolution of both Na+,K(+)-ATPase and AE1, further underlining their potential role in cytoskeleton to membrane linkage.
细胞骨架与膜的关联对于多种细胞功能而言至关重要。红细胞的阴离子交换蛋白（AE1）以及极化上皮细胞的钠钾ATP酶，为整合膜蛋白如何通过连接分子锚蛋白与基于血影蛋白的膜细胞骨架相连，提供了深入研究的范例。在本研究中，我们生成了钠钾ATP酶大的（第三个）胞质结构域（CD3）的多个重组片段，以便在分子水平上确定锚蛋白在CD3上的结合位点。我们提供的证据表明，四个氨基酸组成的簇ALLK对于锚蛋白与重组CD3以及天然钠钾ATP酶的结合至关重要。一旦结合，构象变化可能会揭示钠钾ATP酶上锚蛋白的更多结合位点。与ALLK簇相关的基序也存在于AE1的胞质结构域中，该序列（ALLLK）对锚蛋白结合同样重要。这些基序在钠钾ATP酶和AE1的进化过程中高度保守，进一步突显了它们在细胞骨架与膜连接中的潜在作用。
