Overview

Function of Exchangeable Apolipoproteins
Heart disease is a major cause of mortality in North America. While it is clear that development of cardiovascular disease is a multi-factorial process, it is evident that aberrations in lipid metabolism represent a significant risk factor. It is widely accepted that exchangeable apolipoproteins function in regulation of plasma lipid levels, yet the molecular basis for this role is not fully understood. Increased knowledge of the properties of exchangeable apolipoproteins will be useful in the development of therapeutic strategies to influence plasma lipid levels and thereby, reduce the risk of cardiovascular disease. Furthermore, by understanding the molecular basis of apolipoprotein function in lipid transport and metabolism, it should be possible to design strategies to enhance or interfere with biological processes dependent upon their action. Knowledge gained has direct relevance to the treatment of dyslipidemias, including hypercholesterolemia and disorders of lipid metabolism that affect children and adults.

Ryan Research Program Summary
The laboratory has three main research projects that are focused on a family of plasma proteins that function to regulate lipid transport and metabolism. 

In project 1 the goal is to understand how lipid transport and metabolism are regulated by molecular interactions between lipoproteins and cell surface receptors.  The interaction of apolipoprotein (apo) E with the low-density lipoprotein receptor (LDLR) is under investigation to identify molecular determinants required for a productive receptor-ligand interaction.  We postulated that apoE undergoes a lipid binding induced conformational change that results in extension of helix 4 beyond the boundary identified in its lipid-free helix bundle state (residue 165).  To test the helix extension hypothesis electron paramagnetic spin resonance and NMR spectroscopy analysis of specifically labeled apoE have been performed.  The data reveal that lipid interaction induces a structural transition from random coil (lipid-free state) to alpha helix (lipid associated state) in this region of the protein.  A soluble fragment of LDLR has been employed in ligand binding and release studies with apoE.  LDL-A repeat swapping experiments and modification of the spacer sequences between repeats is under study to determine the molecular requirements for a productive interaction with apoE containing reconstituted high density lipoprotein ligands. 

In project 2 we are exploring the mechanism whereby a recently discovered plasma apolipoprotein, apoA-V, modulates plasma triacylglycerol levels.  Structure-function and site directed mutagenesis experiments have been used to show that apoA-V interacts with heparan sulfate proteglycans (HSPG).  Current research efforts are designed to test the hypothesis that apoA-V functions by facilitating interaction of very low-density lipoproteins (VLDL) with HSPGs, thereby enhancing lipoprotein lipase mediated lipolysis of VLDL associated triacylglycerol.