The overall objective is to define the structure function relationships of human apolipoprotein (apo) E, especially of the isoforms apoE3 and apoE4, to provide a basis for understanding the different cardiovascular disease risks associated with these two proteins. A range of engineered apoE molecules is being used to address 3 specific aims. 1) To determine the secondary structures of human apoE3 and apoE4 in lipid free and lipid bound states using hydrogendeuterium exchange and mass spectrometry methods. The locations and stabilities of 1helical segments in the C-terminal domain of these proteins will be determined to test the hypothesis that these parameters are different in these apoE isoforms. 2) To use a range of physical biochemical methods to identify the region in the C-terminal domain of apoE3 and apoE4 responsible for their different lipid binding properties and lipoprotein binding preferences, and to elucidate the mechanistic basis for these effects. The hypothesis being tested is that the polymorphism alters the structure in the region around residues 260270 so that apoE4 binds better to lipid surfaces. 3) To use adeno-associated viral vectors to express engineered forms of human apoE3 and apoE4 in mice and assess the functional consequences for cholesterol levels and lipoprotein profiles. The hypothesis being tested is that the structure of the C-terminal region spanning residues 260270 controls the different effects that apoE3 and apoE4 have on lipoprotein levels in vivo. Overall, achievement of these 3 aims will generate novel quantitative information about the ways in which apoE structure and polymorphism affect the functional properties of the protein with respect to lipid transport. The design of apoEmimetic molecules and of ways to control the aberrant behavior of apoE4 will be facilitated by this understanding.

Public Health Relevance

In the human population, apoE is expressed in 3 common forms that function differently in regulating lipid transport in vivo. As a result, these mutations in the apoE molecule can increase the risk for development of both cardiovascular disease and Alzheimer's disease. This project will provide more insights into the molecular mechanisms underlying these pathological effects.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL056083-17
Application #
8253719
Study Section
Integrative Nutrition and Metabolic Processes Study Section (INMP)
Program Officer
Liu, Lijuan
Project Start
1997-01-01
Project End
2014-04-30
Budget Start
2012-05-01
Budget End
2013-04-30
Support Year
17
Fiscal Year
2012
Total Cost
$419,352
Indirect Cost
$164,427
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Mizuguchi, Chiharu; Hata, Mami; Dhanasekaran, Padmaja et al. (2014) Fluorescence study of domain structure and lipid interaction of human apolipoproteins E3 and E4. Biochim Biophys Acta 1841:1716-24
Kothapalli, Devashish; Castagnino, Paola; Rader, Daniel J et al. (2013) Apolipoprotein E-mediated cell cycle arrest linked to p27 and the Cox2-dependent repression of miR221/222. Atherosclerosis 227:65-71
Li, Hui; Dhanasekaran, Padmaja; Alexander, Eric T et al. (2013) Molecular mechanisms responsible for the differential effects of apoE3 and apoE4 on plasma lipoprotein-cholesterol levels. Arterioscler Thromb Vasc Biol 33:687-93
Phillips, Michael C (2013) New insights into the determination of HDL structure by apolipoproteins: Thematic review series: high density lipoprotein structure, function, and metabolism. J Lipid Res 54:2034-48
Nguyen, David; Dhanasekaran, Padmaja; Nickel, Margaret et al. (2010) Molecular basis for the differences in lipid and lipoprotein binding properties of human apolipoproteins E3 and E4. Biochemistry 49:10881-9
Lund-Katz, Sissel; Phillips, Michael C (2010) High density lipoprotein structure-function and role in reverse cholesterol transport. Subcell Biochem 51:183-227
Nguyen, David; Dhanasekaran, Padmaja; Phillips, Michael C et al. (2009) Molecular mechanism of apolipoprotein E binding to lipoprotein particles. Biochemistry 48:3025-32
Minagawa, Hirohisa; Gong, Jiang-Sheng; Jung, Cha-Gyun et al. (2009) Mechanism underlying apolipoprotein E (ApoE) isoform-dependent lipid efflux from neural cells in culture. J Neurosci Res 87:2498-508
Sakamoto, Takaaki; Tanaka, Masafumi; Vedhachalam, Charulatha et al. (2008) Contributions of the carboxyl-terminal helical segment to the self-association and lipoprotein preferences of human apolipoprotein E3 and E4 isoforms. Biochemistry 47:2968-77

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