NMR studies of the LDL receptor active conformation of ApoE
Creators: name: University Of California Office Of The President
Publisher: name: University of California Office of the President
publisherIdentifier: https://ror.org/00dmfq477
publisherIdentifierScheme: ROR
Year: 2024

description: Mainstream and environmental tobacco smoke exposure represent significant risk factors for the development of cardiovascular disease in men and women. The relationship between smoking and cardiovascular disease results from multiple mechanisms that interact to contribute to vascular injury and atherosclerotic lesion formation. We are only beginning to understand how smoking contributes to the initiation and progression of cardiovascular disease. One area of intense interest is the effect of tobacco smoke on plasma lipoprotein particles. These particles, which normally function in transport of insoluble lipids through the bloodstream, have been recognized as a target for tobacco combustion-related oxidative damage. A specific type of lipoprotein, termed low density lipoprotein, is adversely affected by oxidation, being converted to a species that accelerates the progression of atherosclerotic lesion development. From knowledge that oxidized low density lipoproteins accumulate at the site of atherosclerotic lesions, it can be concluded that susceptibility of these particles to oxidative modification caused by tobacco smoke exposure may constitute a contributing factor to disease progression. The goal of my research is to understand the molecular basis whereby a protein component of circulating lipoproteins facilitates their removal from the circulation and hence, decreases the amount of low-density lipoprotein available for oxidative modification. This protein component, apolipoprotein E associates with the surface of plasma lipoprotein particles and mediates interaction with cell surface receptor molecules. From knowledge of the structure of apolipoprotein E and the fact that this protein can alter its conformation in such a way that it is, or is not, recognized by lipoprotein receptors forms the basis for our research endeavor. It is known that lipid-association is required to confer this receptor activity to apolipoprotein E. Thus, we hypothesize that increasing the proportion of apolipoprotein E that adopts a receptor-active, lipid-bound state will result in a decrease in plasma low-density lipoprotein concentration and, hence, lowered susceptibility to tobacco smoke related oxidation events and pathological lipoprotein metabolism. In an effort to characterize this lipid bound state that is known to change the conformation of apolipoprotein E, I have devised a strategy to examine a protein region believed to be important in this conformational change in the context of the larger protein. Attempts to examine the entire lipid-bound conformation of the N-terminal domain of apolipoprotein E have proved technically difficult, but using an alternative segmental labeling approach for probing the structure, I hope to circumvent these problems and examine the lipid-induced changes in the protein in a highly specific manner. To accomplish this, I plan to express the protein in two independently generated fragments, one labeled with stable isotopes and another label free. I will then use a protein ligation strategy to join the two protein fragments to construct the intact protein. Once I determine that constructed apolipoprotein E behaves similarly to native apolipoprotein E with respect to folding and lipoprotein receptor activity, I will initiate detailed studies to characterize the lipid-induced structural changes within the labeled domain. By understanding the lipid-induced conformation change within the N-terminal domain of apolipoprotein E at the atomic level, I hope to gain an understanding of how lipid association confers receptor binding activity to the protein. By determining the molecular nature of the receptor active conformation of apolipoprotein E, it will be possible to expand our understanding of the potential for apolipoprotein E to decrease the amount of smoke-induced oxidation-sensitive lipoproteins in plasma.