Cardiovascular disease (CVD) is the number one killer in the United States, taking nearly a million lives each year. It is well established that high density lipoprotein (HDL) and its major protein constituent apolipoprotein (apo) A-l play a major role in reducing the risk of CVD. However the function of apolipoprotein A-l I, the second most abundant protein in HDL, has not been determined. Studies on apoA-ll have led to mixed conclusions concerning the pro- or anti- atherogenicity of apoA-ll in HDL. This study will test the hypothesis that apoA-ll interacts with apoA-l in HDL in a site-specific manner to modulate HDL function. Furthermore, the effect is based on the relative amounts of apoA-l: apoA-ll present in a given HDL particle. In the Mentored Phase of this project, I will determine how the incorporation of apoA-ll affects apoA-l structure in discoidal reconstituted HDL particles. Relative changes in the solvent accessibility of specific apoA-l sequences in mixed particles vs apoA-l only HDL will be assessed using the hydrogen deuterium exchange technique combined with mass spectrometry (HDX-MS). I will then extend the studies in the Independent Phase to locate conformational changes in apoA-l caused by apoA-ll in HDL particles from human plasma. This will be accomplished in stages. First, I will reconstitute spherical HDL particles that resemble native HDL by incorporating varying ratios of apoA-l:apoA-ll along with native HDL lipids. I will also generate native hybrid HDL particles by incorporating isolated apoA-ll into native HDL particles containing apoA-l only. The apoA-ll induced modifications of apoA-l solvent exposure will then be correlated to functional properties including activation of various HDL remodeling factors including lecithin:cholesterol acyl transferase (LCAT), hepatic lipase (HL), and endothelial lipase (EL). I anticipate that this work will provide significant new information on the role of apoA-ll in lipoprotein metabolism and may suggest new therapeutic approaches for fighting CVD.