Stabilization of G protein-coupled receptors by point mutations.
Stabilization of G protein-coupled receptors by point mutations.
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DOI：
10.3389/fphar.2015.00082
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发表时间：
2015
影响因子：
5.6
通讯作者：
Veprintsev DB
中科院分区：
文献类型：
--
作者：
Heydenreich FM;Vuckovic Z;Matkovic M;Veprintsev DB
G蛋白偶联受体（GPCR）是一种参与跨膜信号传导的柔性膜蛋白。它们参与许多生理过程，使它们成为药物开发的有趣目标。确定这些受体的结构将有助于设计更特异的药物，然而，它们的结构表征迄今为止受到低表达和它们在洗涤剂中固有的不稳定性的阻碍，这使得蛋白质工程对于结构和生物物理表征是必不可少的。将受体稳定在特定构象的几种方法已经导致GPCR结构确定的突破。这些包括柔性区域的截短、通过抗体和纳米抗体的稳定化、融合伴侣、高亲和力和共价结合的配体以及通过诱变的构象稳定化。在这篇综述中，我们专注于通过插入点突变来稳定GPCR，从而增加构象和热稳定性以及提高表达水平。我们总结了现有的诱变策略与不同的覆盖范围的GPCR序列空间和深度的信息，设计和突变的可转移性和稳定的分子基础。我们还讨论了突变是否会改变GPCR的结构和药理学特性。
G protein-coupled receptors (GPCRs) are flexible integral membrane proteins involved in transmembrane signaling. Their involvement in many physiological processes makes them interesting targets for drug development. Determination of the structure of these receptors will help to design more specific drugs, however, their structural characterization has so far been hampered by the low expression and their inherent instability in detergents which made protein engineering indispensable for structural and biophysical characterization. Several approaches to stabilize the receptors in a particular conformation have led to breakthroughs in GPCR structure determination. These include truncations of the flexible regions, stabilization by antibodies and nanobodies, fusion partners, high affinity and covalently bound ligands as well as conformational stabilization by mutagenesis. In this review we focus on stabilization of GPCRs by insertion of point mutations, which lead to increased conformational and thermal stability as well as improved expression levels. We summarize existing mutagenesis strategies with different coverage of GPCR sequence space and depth of information, design and transferability of mutations and the molecular basis for stabilization. We also discuss whether mutations alter the structure and pharmacological properties of GPCRs.