Structure and function studies on five homogeneous reconstituted HDL complexes
Hefele Wald, Jennifer Elaine
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Permalink
https://hdl.handle.net/2142/19445
Description
Title
Structure and function studies on five homogeneous reconstituted HDL complexes
Author(s)
Hefele Wald, Jennifer Elaine
Issue Date
1990
Doctoral Committee Chair(s)
Jonas, Ana
Department of Study
Biochemistry
Discipline
Biochemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Chemistry, Biochemistry
Health Sciences, Medicine and Surgery
Language
eng
Abstract
High density lipoprotein (HDL) is believed to be involved in reverse cholesterol transport. HDL consists primarily of phospholipids, cholesterol, cholesterol esters, and apolipoprotein A-I (apo A-I). However, HDL is extremely heterogeneous in size composition and the structure of its lipoprotein components, particularly apo A-I. Due to its inherent structural heterogeneity, structure-function studies on native HDL have been extremely difficult. Techniques were thus developed to reconstitute HDL (rHDL) from apolipoproteins and pure lipids. These rHDL have been heterogeneous in size and the number of apo A-I molecules per complex. This work shows the isolation of 5 rHDL complexes to homogeneity, and the results of studies of rHDL, apo A-I, and lipid structure in these rHDL. Furthermore, the reactivity of these complexes with lecithin cholesterol acyltransferase (LCAT) was also investigated. The rHDL structures were investigated by assays for chemical composition; and the dimensions were determined by native gel electrophoresis and electron microscopy. The protein structure was studied by a variety of fluorescence techniques, and circular dichroism, and infrared spectroscopies. Monoclonal antibody studies, trypsinolysis and structural algorithmic studies were also employed to probe the protein structure. The lipid structure was studied by infrared spectroscopy, and with lipophilic fluorescent probes. Kinetic studies of the complexes were performed with LCAT. This work shows that apo A-I not only has a different structure in the lipid bound versus the lipid free state, but that it has distinct and reproducible structures in the lipid bound state. The alpha helices of apo A-I run parallel to the lipid acyl chains in the discoidal rHDL. The lipid dynamics are affected by both the lipid and protein composition of the complexes; and, differences in rHDL reactivity with LCAT are correlated to differences in lipid content, and apo A-I and rHDL structure.
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