Hulme E C, Bee M S, Goodwin J A
Division of Physical Biochemistry, MRC National Institute for Medical Research, Mill Hill, London NW7 1AA, UK.
Biochem Soc Trans. 2007 Aug;35(Pt 4):742-5. doi: 10.1042/BST0350742.
GPCRs (G-protein-coupled receptors) such as the M(1) muscarinic receptor have so far proved recalcitrant to direct structure determination. Nevertheless systematic mutagenesis, particularly alanine scanning, has advanced our understanding of their structure-function relationships. GPCRs exhibit multiple conformational states with different affinities for and abilities to activate their cognate G-proteins. Ligand binding alters these conformational equilibria, thus promoting or inhibiting signalling. Alanine-scanning mutagenesis probes the relative contributions of a particular amino acid side chain to the stability of the ground and activated states of the receptor and its complexes. These determine the phenotype of the mutant receptor. Classification of the phenotypes suggests functional roles for particular amino acid side chains, allowing us to group them accordingly. From a rhodopsin-based homology model of the M(1) mAChR, a coherent view emerges of how these clusters of residues function in ligand anchoring, transduction of binding energy, global structural stabilization and selective stabilization of the ground state or the activated state of the receptor. We can identify differences in ligand-binding modes, and suggest inter- and intra-molecular interactions that are weakened or broken, or formed or intensified during acetylcholine-induced activation. In due course, we may be able to extend these insights to activation by unconventional agonists.
到目前为止，诸如M(1)毒蕈碱受体之类的G蛋白偶联受体（GPCRs）难以直接进行结构测定。然而，系统诱变，尤其是丙氨酸扫描，增进了我们对其结构-功能关系的理解。GPCRs呈现出多种构象状态，对其同源G蛋白具有不同的亲和力和激活能力。配体结合会改变这些构象平衡，从而促进或抑制信号传导。丙氨酸扫描诱变可探究特定氨基酸侧链对受体及其复合物基态和激活态稳定性的相对贡献。这些因素决定了突变受体的表型。对表型进行分类可揭示特定氨基酸侧链的功能作用，从而使我们能够据此对它们进行分组。从基于视紫红质的M(1)毒蕈碱型乙酰胆碱受体同源模型中，可以清晰地看出这些残基簇在配体锚定、结合能转导、整体结构稳定以及受体基态或激活态的选择性稳定中是如何发挥作用的。我们能够识别配体结合模式的差异，并提出在乙酰胆碱诱导的激活过程中被削弱或破坏、或形成或增强的分子间和分子内相互作用。假以时日，我们或许能够将这些见解扩展至非传统激动剂引发的激活作用。
