Solution NMR approaches for establishing specificity of weak heterodimerization of membrane proteins.
Solution NMR approaches for establishing specificity of weak heterodimerization of membrane proteins.
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溶液核磁共振方法用于确定膜蛋白弱异二聚化的特异性。
DOI：
10.1021/ja208972h
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发表时间：
2011
影响因子：
15
通讯作者：
Sanders,CharlesR
中科院分区：
文献类型：
--
作者：
Zhuang,Tiandi;Jap,BingK;Sanders,CharlesR
溶液核磁共振提供了一个强有力的方法来检测复杂的形成涉及弱到中等的分子间的亲和力。然而，溶液NMR很少用于检测模型膜中两种膜蛋白之间的复合物形成。特异性结合对膜蛋白NMR谱的影响很难与非特异性结合引起的光谱变化和/或两种蛋白在单一模型膜组件中强制共存引起的变化区分开来。当溶解度限制使得不可能完成滴定至络合物形成的接近饱和点时，情况尤其如此。在这项工作中，实验提供的基础上，建立是否发生特定的复合物形成两个膜蛋白之间的条件下，结合是不是高亲和力。这些方法的应用导致以下结论：膜蛋白CD 147（也称为EMMPRIN或basigin）在膜中与淀粉样前体蛋白（C99或APP-βCTF）的99个残基跨膜C-末端片段形成特异性异二聚体复合物，后者是与阿尔茨海默病病因学密切相关的淀粉样β多肽的直接前体。
Solution NMR provides a powerful approach for detecting complex formation involving weak to moderate intermolecular affinity. However, solution NMR has only rarely been used to detect complex formation between two membrane proteins in model membranes. The impact ofspecific bindingon the NMR spectrum of a membrane protein can be difficult to distinguish from spectral changes that are induced bynonspecific bindingand/or by changes that arise fromforced cohabitationof the two proteins in a single model membrane assembly. This is particularly the case when solubility limits make it impossible to complete a titration to the point of near saturation of complex formation. In this work experiments are presented that provide the basis for establishing whether specific complex formation occurs between two membrane proteins under conditions where binding is not of high avidity. Application of these methods led to the conclusion that the membrane protein CD147 (also known as EMMPRIN or basigin) forms a specific heterodimeric complex in the membrane with the 99-residue transmembrane C-terminal fragment of the amyloid precursor protein (C99 or APP-βCTF), the latter being the immediate precursor of the amyloid-β polypeptides that are closely linked to the etiology of Alzheimer’s disease.