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 relativ...

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Veröffentlicht in:The Journal of biological chemistry 2006-07, Vol.281 (29), p.20418-20426
Hauptverfasser: Martin, Dale D.O., Budamagunta, Madhu S., Ryan, Robert O., Voss, John C., Oda, Michael N.
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container_end_page 20426
container_issue 29
container_start_page 20418
container_title The Journal of biological chemistry
container_volume 281
creator Martin, Dale D.O.
Budamagunta, Madhu S.
Ryan, Robert O.
Voss, John C.
Oda, Michael N.
description 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.
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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. 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source MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; PubMed Central; Alma/SFX Local Collection
subjects Apolipoprotein A-I - chemistry
Apolipoprotein A-I - genetics
DNA Primers
Electron Spin Resonance Spectroscopy
Fluorescence Resonance Energy Transfer
Genetic Variation
Humans
Lipoproteins, HDL - chemistry
Models, Molecular
Mutagenesis, Site-Directed
Protein Conformation
Recombinant Proteins - chemistry
title Apolipoprotein A-I Assumes a “Looped Belt” Conformation on Reconstituted High Density Lipoprotein
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