Structure, Dynamics and Activation Mechanisms of Chemokine Receptors

描述（由申请人提供）：G蛋白偶联受体（gpcr）是人类最大的蛋白质超家族，有近1000个成员。它们是7种跨膜受体，通过广泛的刺激转导来协调细胞间的通讯，这些刺激涉及感觉、神经传递、发育、情感、认知以及中枢神经系统、内分泌和免疫系统的功能。趋化因子受体是一类重要的gpcr，以其在免疫监视中的关键作用而闻名，它们控制白细胞的迁移和激活，以检测和解决诸如癌症和感染等生理异常。然而，这些受体的不适当表达或调节与包括炎症性疾病、癌症和艾滋病在内的大量病理有关；因此，开发阻断特定趋化因子受体功能的小分子受体拮抗剂具有重要意义。直到最近，由于受体表达和结晶方面的挑战，gpcr一直无法确定其结构。然而，新技术的出现使得确定GPCR结构的可行性无可争议。为此，我们的主要目标是获得趋化因子受体和受体复合物的结构信息，这些信息可以帮助药物发现工作，旨在提高亲和力，有效性和选择性。因此，我们将与PSI网络合作，提出申请

DESCRIPTION (provided by applicant): G protein coupled receptors (GPCRs) represent the largest protein superfamily in humans, with nearly 1000 members. They are seven transmembrane receptors that coordinate intercellular communication via the transduction of a wide range of stimuli involved in sensation, neurotransmission, development, emotion, cognition, and function in the CNS, endocrine and immune systems. Chemokine receptors are an important class of GPCRs that are best known for their pivotal role in immune surveillance, where they control the migration and activation of leukocytes in an effort to detect and resolve physiological abnormalities such as cancer and infection. However, inappropriate expression or regulation of these receptors is associated with an extraordinary number of pathologies including inflammatory diseases, cancer and AIDS; thus there is significant interest in developing small molecule receptor antagonists that block the function of specific chemokine receptors. Until recently, GPCRs had eluded structure determination due to challenges in receptor expression and crystallization. However, new technologies have emerged which has made the viability of determining GPCR structures indisputable. To this end, our primary goal is to obtain structural information on chemokine receptors and receptor complexes that can aid drug discovery efforts aimed at improving affinity, efficacy, and selectivity. Accordingly, in collaboration with the PSI network, we will apply novel technologies for the expression, purification and crystallization of GPCRs from the chemokine receptor family, with the goal of determining at least two different receptor structures and multiple co-complexes by the five-year endpoint. To maximize the capabilities of the PSI centers in generating purified protein, and to acquire insights into the dynamic aspects of receptor function, the crystallographic work will be complemented with biophysical studies. Radiolytic footprinting will be developed and applied to map the binding interfaces between chemokines and receptors and to determine information on activation mechanisms. The interaction of pathogenic proteins with chemokine receptors will also be investigated, specifically, the CCR5 receptor with the HIV glycoprotein gp120, and the DARC receptor with the malarial docking protein, DBP. Site Directed Spin Labeling with Electron Paramagnetic Resonance (SDSL-EPR) will be used to characterize the conformational changes associated with ligand binding. All of these studies will be augmented with computational modeling methods in order to rationally guide the experimental construct design and to interpret the biophysical data in a 3D context.