G-蛋白偶联受体的激活构象构建与激活机制的理论研究

G-protein coupled receptor (GPCR) is one of the indispensable protein families in human body. There are about 800 genes encoding GPCR in human. As a membrane protein, GPCR can sense the activation signal from the extracellular side of the cell and transfer the signal to the intracellular side of the cell to regulate versatile physiological behaviors of the cell. Thus, altering the activation of the GPCR can be an effective way to regulate the dysfunction of the cell, and there are about 30% of the clinic used drugs targeting GPCR. Although with a diverse of functions, the structures of GPCR are conserved, which is characterized by seven transmembrane helixes. Such a seven-transmembrane helix structure was first resolved in 2000 by Palczewski and his coworkers. In the following years, GPCR community has experienced a flurry of GPCR structures. By now, there are 170 GPCR crystal structures belonging to 35 distinct GPCR members are resolved. These structures provide us with atomic resolutions of the protein. On the other hand, most of the proteins are crystallized in the inactive state. To the limit of our knowledge, only 6 GPCR structures (rhodopsin, adenosine A2A receptor, β2-adrenergic receptor, M2 muscarinic receptor, μ-opioid receptor, and viral US28) in the active state are resolved. Because of the lacking GPCR structures in the active state, the studies of GPCR activation mechanism is greatly hindered. In this proposal, we would like to employ computation methods, including molecular dynamics simulations, metadynamics simulations, and Markov state models, to investigate the activation mechanism of GPCR.

G-蛋白偶联受体是人体中最重要的膜蛋白家族之一。它包含近800名成员，这些成员能够接受细胞膜外刺激，将这些信号传递到细胞内部，以调节细胞各种生理活动。鉴于此类蛋白在信号传递中的重要作用，临床上约有30%的药物都作用于它。尽管功能多样，但在结构上，它们却有保守的七跨膜螺旋的结构。这种结构特征在2000年发表的第一个高分辨率G-蛋白偶联受体（rhodopsin）结构中得到确认。 随后，随着新技术的应用，特别是lipid cubic 技术的引入，大大提高了结构解析的成功率。自2007以来，又有170多个分布于35个G-蛋白偶联受体的结构得到解析。但激活状态下的受体比其在抑制状态更加不稳定，所以在已知结构中，只有六个结构处于激活构象。这大大限制了我们对G-蛋白偶联受体的作用机制的研究。鉴于此，我们希望通过计算机模拟的方法构建G-蛋白偶联受体的激活构象，并对其作用机制进行研究。

胰高血糖素样肽-1受体（GLP-1R）属于B类GPCR，是治疗II型糖尿病和肥胖的靶标。目前，获批用于治疗II型糖尿病的几个多肽类药物，除口服索马鲁肽外，均是注射剂型，且会带来如恶心和呕吐等副作用。口服非多肽类药物一直是研究机构和工业界尝试用于治疗II型糖尿病及减肥的药物，但由于小分子难以模拟受体与多肽的相互作用，限制了胰高血糖素样肽-1受体小分子药物的发现。本项目测定了小分子完全激动剂RGT1383结合GLP-1R并与Gs蛋白复合的结构。与已有报道的激动剂结合的GLP-1R结构相比，本项目最大的发现是非肽类激动剂RGT1383由于ECL3区域和TM7区域的胞外末端的向内运动而结合到受体的顺式结合口袋。另外，NTD区域的W33在RGT1383与受体的结合中起到了关键作用，解释了NTD调节的GLP-1R激活机制并指明了这一类小分子激动剂的种属特异性。本项目对设计口服非肽类小分子GLP-1R激动剂类药物用于治疗二型糖尿病和肥胖具有一定的指导意义。