PROJECT 2: STRUCTURAL BASIS OF THE ANTI-ATHEROGENIC PROPERTIES OF APO A-l The goal of this project is to elucidate the molecular mechanisms underlying the functions of apolipoprotein (apo) A-l in reverse cholesterol transport (RCT). ApoA-l is the major protein of plasma highdensity lipoprotein (HDL) and the functions of this molecule underlie the anti-atherogenic properties of the lipoprotein.
Specific Aim 1 is to define the key properties of the two tertiary structure domains of the apoA-l molecule, and to better define lipid-free human apoA-l structure in dilute solution at physiological concentrations. The influence of the properties of the two apoA-l structural domains on the affinity and mechanism of lipid binding will also be investigated. These objectives will be accomplished by using engineered apoA-l molecules and a range of physical-biochemical methods.
Specific Aim 2 is to establish the mechanistic basis for the biogenesis of heterogeneous nascent HDL particles via the ATP-binding cassette transporter Al (ABCA1 )-mediated efflux of cellular lipids to apoA-l. The role of apoA-l structure and influence of cell type will be determined by measuring the efflux of phospholipid and cholesterol molecules from macrophages, fibroblasts and liver cells growing in culture to engineered apoA-l molecules.
Specific Aim 3 is to define the effects of the properties of the tertiary structural domains of apoA-l on the protein's functionality in cholesterol transport using adeno-associated virus-induced expression of natural human apoA-l variants in mice to assess the effects on RCT and atherosclerosis (in collaboration with Project 3). The functionalities of the mouse HDL particles containing the apoA-l mutations in mediating cholesterol transport into and out of cells will be determined in collaboration with Project 1. The incidence of premature coronary artery disease is reduced in human populations with elevated levels of plasma HDL cholesterol and apoA-l. The reasons for this protective effect are not understood fully and this project seeks to uncover the molecular mechanisms underlying the beneficial properties of apoA-l.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Program Projects (P01)
Project #
5P01HL022633-35
Application #
8374991
Study Section
Heart, Lung, and Blood Initial Review Group (HLBP)
Project Start
Project End
Budget Start
2011-12-01
Budget End
2012-11-30
Support Year
35
Fiscal Year
2012
Total Cost
$305,340
Indirect Cost
$94,804
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Chetty, Palaniappan Sevugan; Nguyen, David; Nickel, Margaret et al. (2013) Comparison of apoA-I helical structure and stability in discoidal and spherical HDL particles by HX and mass spectrometry. J Lipid Res 54:1589-97
Nguyen, David; Nickel, Margaret; Mizuguchi, Chiharu et al. (2013) Interactions of apolipoprotein A-I with high-density lipoprotein particles. Biochemistry 52:1963-72
Alexander, Eric T; Phillips, Michael C (2013) Influence of apolipoprotein A-I and apolipoprotein A-II availability on nascent HDL heterogeneity. J Lipid Res 54:3464-70
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

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