The major objectives of this project are to provide detailed structural, conformational and dynamic information on the molecular organization and interactions of lipids and apoproteins in the plasma lipoproteins (primarily HDL and LDL). Studies focus on high resolution structural investigations of specific apoproteins and lipoproteins using state-of- the-art methodologies of structural biology. The primary areas explored concern: a. the three dimensional structure of intact lipoproteins (HDL, LDL) with emphasis on the molecular conformation of the apoproteins of the lipoprotein surface, and b. the detailed molecular conformation and structure of the apolipoproteins (AI and B). Studies of apolipoproteins AI will employ a combination of methodologies for the identification of important structural and functional elements in the sequence. Synthetic peptides will be used to model these regions. The detailed molecular conformation and thermodynamic stability will be studied by modelling, X- ray crystallography, 2D NMR, CD, and calorimetry. In addition the detailed molecular structure of the native apoproteins will be studied by X-ray crystallography. Studies of apoprotein B will focus on the domain folding of the protein and the conformation and function of the subdomains. Protease cleavage and molecular biology approaches will be used to obtain segments of the apoB sequence representing these structural domains. Structural studies of native LDL, by cryo electron microscopy will focus on the organization of apoB and the localization of structural and functional domains on the LDL particle using a combination of immuno- nanogold labelling. A detailed molecular understanding of apoprotein conformation and structure is vital to further progress in understanding lipoprotein structure and function. The information derived in this project will result in an improved understanding of the molecular organization and interactions in HDL and LDL. More importantly, they will provide basic background information essential to understanding physiological and metabolic processes such as lipoprotein cell surface interactions, lipoprotein interconversion and catabolism at a truly molecular level.
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