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J Lipid Res. 2018 Jul;59(7):1244-1255. doi: 10.1194/jlr.M085332. Epub 2018 May 17.

A thumbwheel mechanism for APOA1 activation of LCAT activity in HDL.

Author information

1
Departments of Pathology and Laboratory Medicine University of Cincinnati, Cincinnati, OH 45237.
2
Departments of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, TN 37232.
3
Division of Immunobiology and Center for Systems Immunology Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229.
4
Anesthesiology, Vanderbilt University Medical Center, Nashville, TN 37232.
5
Medicine, Vanderbilt University Medical Center, Nashville, TN 37232.
6
Lipoprotein Metabolism Section, Cardiovascular-Pulmonary Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.
7
Division of Endocrinology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229.
8
Molecular Genetics, Biochemistry, and Microbiology, University of Cincinnati, Cincinnati, OH 45237.
9
Departments of Pathology and Laboratory Medicine University of Cincinnati, Cincinnati, OH 45237 Sean.Davidson@UC.edu.

Abstract

APOA1 is the most abundant protein in HDL. It modulates interactions that affect HDL's cardioprotective functions, in part via its activation of the enzyme, LCAT. On nascent discoidal HDL, APOA1 comprises 10 α-helical repeats arranged in an anti-parallel stacked-ring structure that encapsulates a lipid bilayer. Previous chemical cross-linking studies suggested that these APOA1 rings can adopt at least two different orientations, or registries, with respect to each other; however, the functional impact of these structural changes is unknown. Here, we placed cysteine residues at locations predicted to form disulfide bonds in each orientation and then measured APOA1's ability to adopt the two registries during HDL particle formation. We found that most APOA1 oriented with the fifth helix of one molecule across from fifth helix of the other (5/5 helical registry), but a fraction adopted a 5/2 registry. Engineered HDLs that were locked in 5/5 or 5/2 registries by disulfide bonds equally promoted cholesterol efflux from macrophages, indicating functional particles. However, unlike the 5/5 registry or the WT, the 5/2 registry impaired LCAT cholesteryl esterification activity (P < 0.001), despite LCAT binding equally to all particles. Chemical cross-linking studies suggest that full LCAT activity requires a hybrid epitope composed of helices 5-7 on one APOA1 molecule and helices 3-4 on the other. Thus, APOA1 may use a reciprocating thumbwheel-like mechanism to activate HDL-remodeling proteins.

KEYWORDS:

apolipoprotein A1; apolipoproteins; cholesterol metabolism; cholesterol/efflux; electron microscopy; high density lipoprotein; high density lipoprotein metabolism; lecithin:cholesterol acyltransferase; proteomics; surface plasmon resonance

PMID:
29773713
PMCID:
PMC6027914
[Available on 2019-07-01]
DOI:
10.1194/jlr.M085332

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