Class B, type I scavenger receptor (SR-BI) can bind high density lipoprotein (HDL) and mediate HDL selective lipid uptake. Genetic and physiological studies suggest that SR-BI's function may be directly linked to health and diseases, e.g., atherosclerosis. However,, the mechanism of HDL selective lipid uptake remains poorly understood. Therefore, this research proposal addresses the fundamental question: what is the mechanism of SR-BI-mediated selective lipid uptake? The specific aims of this work are. First, structure and function analysis of SR-BI: 1) Can SR-BI work alone in mediating selective uptake or does it need protein co-factors? 2) If it does, the goal is to identify these co- factors. 3) Does the efficiency of SR-BI-mediated selective uptake depend on the membrane lipid composition depend on the membrane lipid composition? 4) Does N-linked glycosylation play a role in SR-BI- mediated selective lipid uptake? 5) Crystallization of SR-BI- and SR-BI mutants. Second, monoclonal antibody preparation for SR-BI localization studies on physiological effects of blocking SR-BI's function, and co- immunoprecipitation (IP) of SR-BI. The techniques that will be used for this work include site-directed mutagenesis, cross-linking, co-IP, in vitro reconstitution, fluorescence activated cell sorting (FACS), antibody affinity chromatography, enzyme-linked immunosorbent assay (ELISA) and immunoelectron microscopy. This work is critical in elucidating the mechanism of SR-BI mediated selective uptake. Therefore, the result of this research can provide a tool for in vitro functional analysis of SR-BI , and may serve as the guideline for designing future drugs to treatment and/or prevent atherosclerosis.
