A mass spectrometric determination of the conformation of dimeric apolipoprotein A-I in discoidal high density lipoproteins.
Discoidal forms of high density lipoproteins (HDL) are critical intermediates between lipid-poor apolipoprotein A-I (apo A-I), the major protein constituent of HDL, and the mature spherical forms that comprise the bulk of circulating particles. Thus, many studies have focused on understanding apoA-I structure in discs reconstituted in vitro. Recent theoretical and experimental work supports a "belt" model for apoA-I in which repeating amphipathic helical domains run parallel to the plane of the lipid disc. However, disc-associated apoA-I can adopt several tertiary arrangements that are consistent with a belt orientation. To distinguish among these, we cross-linked near-neighbor Lys groups in homogeneous 96 A discs containing exactly two molecules of apoA-I. After delipidation and tryptic digestion, mass spectrometry was used to identify 9 intermolecular and 11 intramolecular cross-links. The cross-linking pattern strongly suggests a "double-belt" molecular arrangement for apoA-I in which two apoA-I molecules wrap around the lipid bilayer disc forming two stacked rings in an antiparallel orientation with helix 5 of each apoA-I in juxtaposition (LL5/5 orientation). The data also suggests the presence of an additional double-belt orientation with a shifted helical registry (LL5/2 orientation). Furthermore, a 78 A particle with two molecules of apoA-I fit a similar double-belt motif with evidence for conformational changes in the N-terminus and the region near helix 5. A comparison of this work to a previous study is suggestive that a third molecule of apoA-I can form a hairpin in larger particles containing three molecules of apoA-I.
Silva
R A Gangani D
RA
Department of Pathology and Laboratory Medicine, The University of Cincinnati, Cincinnati, Ohio 45237-0507, USA.
Hilliard
George M
GM
Li
Ling
L
Segrest
Jere P
JP
Davidson
W Sean
WS
D000595:Amino Acid Sequence; D016632:Apolipoprotein A-I / Q000737:chemistry*; D019281:Dimerization; D006801:Humans; D008075:Lipoproteins, HDL / Q000737:chemistry*; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D017433:Protein Structure, Secondary
D016632:Apolipoprotein A-I; D008075:Lipoproteins, HDL; D010446:Peptide Fragments