Apolipoprotein A-I assumes a "looped belt" conformation on reconstituted high density lipoprotein.
Apolipoprotein A-I (apoA-I) plays a central role in the reverse cholesterol transport pathway; however, the structural basis for its antiatherogenic effects remains poorly understood. Here we employ EPR spectroscopy and fluorescence resonance energy transfer to elucidate the conformation and relative alignment of apoA-I monomers on discoidal (9.4 nm) reconstituted high density lipoprotein (rHDL). EPR spectroscopy provided evidence for an extended helical secondary structure. Position 139 since it was the only residue examined to display a dynamic motional character consistent with a flexible loop structure. The EPR spectra of nitroxide probes at positions 133 and 146 exhibit spin coupling, indicating that these positions are proximal to an apoA-I paired counterpart on the perimeter of rHDL. fluorescence resonance energy transfer studies employing engineered apoA-I variants possessing a single tryptophan (energy donor) and/or a single cysteine (whose thiol moiety was covalently labeled with an extrinsic energy acceptor) provided evidence that paired apoA-I molecules around the perimeter of rHDL align in an extended antiparallel conformation. Taken together with the observation that the EPR spectra of nitroxide probes positioned at intervening sequence positions (134-145) do not exhibit spin coupling, this has led us to propose a "looped belt" model, wherein residues 133-146 comprise a flexible loop segment that confers to apoA-I an intrinsic ability to adapt its structure to accommodate changing particle lipid content. Specifically, in the looped belt model, with the exception of amino acids 134-145, apoA-I aligns with its counterpart in a helix 5-helix 5 registry, centered at position 139.
Martin
Dale D O
DD
Children's Hospital Oakland Research Institute, Oakland, California 94609-1673, USA.
Budamagunta
Madhu S
MS
Ryan
Robert O
RO
Voss
John C
JC
Oda
Michael N
MN
D016632:Apolipoprotein A-I / Q000737:chemistry* / Q000235:genetics; D017931:DNA Primers; D004578:Electron Spin Resonance Spectroscopy; D031541:Fluorescence Resonance Energy Transfer; D014644:Genetic Variation; D006801:Humans; D008075:Lipoproteins, HDL / Q000737:chemistry*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry
D016632:Apolipoprotein A-I; D017931:DNA Primers; D008075:Lipoproteins, HDL; D011994:Recombinant Proteins