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.
