STRUCTURAL BASIS OF CHEMOKINE RECEPTOR RECOGNITION

趋化因子的生物学功能主要依赖于它们对G蛋白偶联受体(GPCRs)的识别和激活。拟议项目的总体目标是了解趋化因子及其受体之间的结构相互作用。不幸的是，由于处理全长GPCR的困难，X射线结晶学或核磁共振(核磁共振)光谱无法直接观察到这些相互作用。突变研究已经确定了参与功能性蛋白质-蛋白质相互作用的残基，但导致受体识别和激活的确切相互作用仍未确定。在拟议的研究中采用的方法试图通过使用来自趋化因子受体胞外区域的合成肽来重建识别界面来绕过这一问题。这些基于受体的多肽的构象以及它们与溶液中趋化因子的相互作用将用圆二色谱和核磁共振光谱来表征。这些多肽的生物活性将通过体外中性粒细胞趋化和受体结合试验来确定。将对这些数据进行分析，以开发趋化因子受体识别的结构模型，该模型将使用定点突变进行测试，然后进行功能分析。趋化因子受体识别的结构基础研究与人类健康和疾病有着广泛的相关性。趋化因子在包括哮喘、关节炎和牛皮癣在内的各种过敏性和风湿性疾病中起主要作用。趋化因子在动脉粥样硬化、血管生成和肿瘤生长中的作用已有报道。一些趋化因子受体作为HIV-1辅助受体发挥作用，已成为识别新的抗HIV化合物的有吸引力的靶点。受体识别模型将指导实验描绘特定趋化因子-受体相互作用的生物学作用。此外，治疗剂可以被合理地设计成精确地抑制与疾病有特殊关系的单对趋化因子及其受体。拟议中的项目将在Elias Lolis博士的实验室进行，这将使我能够继续接受蛋白质结构生物学方面的培训，并向我介绍免疫生物学的科学和技术。通过这次培训，我的职业发展将得到加强，并为我未来在学术医学研究领域的职业生涯做好准备。

The biological function of chemokines is critically dependent on their recognition and activation of specific G protein coupled receptors (GPCRs). The overall goal of the proposed project is to develop an understanding of the structural interactions between chemokines and their receptors. Unfortunately, these interactions cannot be directly observed by X-ray crystallography or nuclear magnetic resonance (NMR) spectroscopy due to the difficulty of working with full-length GPCRs. Mutagenesis studies have identified residues that are involved in functional protein-protein interactions, but the precise interactions that result in receptor recognition and activation remain uncharacterized. The approach adopted in the proposed research attempts to circumvent this problem by reconstructing the recognition interface using synthetic peptides derived from the extracellular domains of chemokine receptors. The conformation of these receptor-based peptides and their interactions with chemokines in solution will then be characterized using circular dichroism and NMR spectroscopy. The biological activity of the peptides will be determined using in vitro assays of neutrophil chemotaxis and receptor binding. The data will be analyzed in order to develop a structural model of chemokine receptor recognition which will be tested using site-directed mutagenesis, followed by functional assays. Research into the structural basis of chemokine receptor recognition has wide-ranging relevance to human health and disease. Chemokines play a predominant role in a variety of allergic and rheumatic diseases including asthma, arthritis and psoriasis. Effects of chemokines on atherogenesis, angiogenesis and tumor growth have been reported. Some of the chemokine receptors function as HIV-1 coreceptors and have become attractive targets for identifying new anti-HIV compounds. A model of receptor recognition will guide experiments to delineate the biological roles of specific chemokine-receptor interactions. As well, therapeutic agents could be rationally designed to precisely inhibit single pairs of chemokines and their receptors which are specifically implicated in disease. The proposed project to be performed in the laboratory of Dr. Elias Lolis will allow me to continue my training in protein structural biology as well as introduce me to the science and techniques of immunobiology. My career development will be enhanced by this training and will prepare me for a future career in academic medical research.