High density lipoprotein (HDL) and its major protein constituent, apolipoprotein (apo)A-l, play critical roles in the prevention of human cardiovascular disease, which claims nearly a million lives in the United States every year. Unfortunately, there are still many unanswered questions about the molecular basis for the cardio-protective effects of HDL. A prominent obstacle in the way of a detailed understanding of these effects is the lack of information on the structure of apoA-l in HDL particles that exist in human plasma. We propose to test the hypothesis that the structure of apoA-l in spherical plasma HDL particles is related to that found in simple HDL particles that can be created and studied in detail in vitro. In the initial funding period, we used cross-linking chemistry and high-resolution mass spectrometry to generate detailed models of apoA-l in reconstituted particles of various size and shape. In this continuation application, we propose to extend these models to real HDL particles isolated from human plasma. Using the structures we have already worked out, we will monitor the conformation of apoA-l as the complexity of the particles is systematically increased. As a first step, the effects of introducing apolipoprotein A-ll, the second most common protein constituent of HDL, on the structure of apoA-l will be evaluated. We will then monitor the structure of apoA-l and A-ll in spherical particles generated with lipid fractions isolated from plasma HDL. Finally, true human plasma particles will be used to test our models. We believe that this will be the most comprehensive study of apoA-l and apoA-ll structure in authentic human HDL particles ever attempted. We will also study the functional consequences of a shift in apoA-l molecular registry in discoidal and spherical complexes that may have exciting implications for how HDL composition modulates its metabolism. It is clear that the nature of plasma HDL precludes the use of NMR and X-ray crystallography for detailed structural studies, at least with current technology. However, our mass spectrometry approach is not limited by the requirement of a homogeneous sample and is thus the best available for addressing this important and complex problem. We believe that the information from these studies will provide the basis for future highly targeted mutagenesis strategies designed to dissect out the protective functions of apoA-l, perhaps leading to therapies for enhancing the protective effects of HDL.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL067093-07
Application #
7624190
Study Section
Integrative Nutrition and Metabolic Processes Study Section (INMP)
Program Officer
Liu, Lijuan
Project Start
2001-04-01
Project End
2012-05-31
Budget Start
2009-06-01
Budget End
2010-05-31
Support Year
7
Fiscal Year
2009
Total Cost
$347,639
Indirect Cost
Name
University of Cincinnati
Department
Pathology
Type
Schools of Medicine
DUNS #
041064767
City
Cincinnati
State
OH
Country
United States
Zip Code
45221
Swertfeger, Debi K; Li, Hailong; Rebholz, Sandra et al. (2017) Mapping Atheroprotective Functions and Related Proteins/Lipoproteins in Size Fractionated Human Plasma. Mol Cell Proteomics 16:680-693
Melchior, John T; Street, Scott E; Andraski, Allison B et al. (2017) Apolipoprotein A-II alters the proteome of human lipoproteins and enhances cholesterol efflux from ABCA1. J Lipid Res 58:1374-1385
Davidson, W Sean; Heink, Anna; Sexmith, Hannah et al. (2017) Obesity is associated with an altered HDL subspecies profile among adolescents with metabolic disease. J Lipid Res 58:1916-1923
Davidson, W S; Inge, T H; Sexmith, H et al. (2017) Weight loss surgery in adolescents corrects high-density lipoprotein subspecies and their function. Int J Obes (Lond) 41:83-89
Melchior, John T; Walker, Ryan G; Morris, Jamie et al. (2016) An Evaluation of the Crystal Structure of C-terminal Truncated Apolipoprotein A-I in Solution Reveals Structural Dynamics Related to Lipid Binding. J Biol Chem 291:5439-51
Davidson, W Sean; Heink, Anna; Sexmith, Hannah et al. (2016) The effects of apolipoprotein B depletion on HDL subspecies composition and function. J Lipid Res 57:674-86
Gordon, Scott M; Li, Hailong; Zhu, Xiaoting et al. (2016) Impact of genetic deletion of platform apolipoproteins on the size distribution of the murine lipoproteome. J Proteomics 146:184-94
Unruh, Dusten; Srinivasan, Ramprasad; Benson, Tyler et al. (2015) Red Blood Cell Dysfunction Induced by High-Fat Diet: Potential Implications for Obesity-Related Atherosclerosis. Circulation 132:1898-908
Gordon, Scott M; Li, Hailong; Zhu, Xiaoting et al. (2015) A comparison of the mouse and human lipoproteome: suitability of the mouse model for studies of human lipoproteins. J Proteome Res 14:2686-95
Heink, Anna; Davidson, W Sean; Swertfeger, Debi K et al. (2015) A Comparison of Methods To Enhance Protein Detection of Lipoproteins by Mass Spectrometry. J Proteome Res 14:2943-50

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