Structure and function studies on five homogeneous reconstituted HDL complexes

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.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biochemistry

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2011

Author dc:creator

• Hefele Wald, Jennifer Elaine

Contributors dc:contributor

• Jonas, Ana

• Chemistry, Biochemistry
• Health Sciences, Medicine and Surgery

Statement dc:rights

• Copyright 1990 Hefele Wald, Jennifer Elaine

Language dc:language

eng

Identifier

AAI9114262
(UMI)AAI9114262

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/19445