Defined apolipoprotein A-I conformations in reconstituted high density lipoprotein discs.
We prepared and isolated defined, reconstituted high density lipoprotein (r-HDL) particles containing apolipoprotein A-I (apoA-I), palmitoyloleoylphosphatidylcholine, and cholesterol. The initial r-HDL were prepared by the sodium cholate method, then part of the preparation was depleted of phospholipid by exposure to LDL, and the resulting, stable r-HDL species were isolated by gel filtration. The isolated r-HDL were characterized in terms of their size, alpha-helix content, and the conformation of apoA-I as reported by the fluorescence properties of the tryptophan residues. Then the relative reactivity of the r-HDL with lecithin cholesterol acyltransferase was assessed. The isolated, discoidal r-HDL contained 2 and 3 apoA-I molecules/particle, and had 77 and 109 A diameters, respectively. Their spectral properties were essentially identical and were distinct from the larger particles in the class of r-HDL with 2 apoA-I molecules/particle (particles with diameters of 86 and 96 A). In addition, the reactivity of the 77 and 109 A particles with pure lecithin cholesterol acyltransferase was similar and about 10-fold lower than for the 86 and 96 A particles. We conclude that the stable, limiting r-HDL particles in each class (77 and 109 A) can arise from the larger particles of the same class by depletion of phospholipids. These limiting particles have very similar apoA-I conformations, with decreased alpha-helix contents and compact protein regions, that are very poor in activating lecithin cholesterol acyltransferase. Based on these results, we propose a model to explain the origin of the different classes and subclasses of the discoidal r-HDL particles.
Jonas
A
A
Department of Biochemistry, College of Medicine at Urbana-Champaign, Illinois 61801.
Kézdy
K E
KE
Wald
J H
JH
D016632:Apolipoprotein A-I; D001054:Apolipoproteins A / Q000302:isolation & purification; D002784:Cholesterol / Q000302:isolation & purification; D003461:Crystallography; D003720:Densitometry; D005454:Fluorescence Polarization; D006801:Humans; D008075:Lipoproteins, HDL / Q000302:isolation & purification; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy; D007862:Phosphatidylcholine-Sterol O-Acyltransferase / Q000378:metabolism; D011487:Protein Conformation; D013379:Substrate Specificity
D016632:Apolipoprotein A-I; D001054:Apolipoproteins A; D008075:Lipoproteins, HDL; D046911:Macromolecular Substances; D002784:Cholesterol; D007862:Phosphatidylcholine-Sterol O-Acyltransferase