title: Membrane-bound alpha-synuclein forms an extended helix: long-distance pulsed ESR measurements using vesicles, bicelles, and rodlike micelles.
abstract: We apply pulsed dipolar ESR spectroscopy (Ku-band DEER) to elucidate the global conformation of the Parkinson's disease-associated protein, alpha-synuclein (alphaS) bound to small unilamellar phospholipid vesicles, rodlike SDS micelles, or lipid bicelles. By measuring distances as long as approximately 7 nm between introduced pairs of nitroxide spin labels, we show that distances are close to the expectations for a single continuous helix in all cases studied. In particular, we find distances of 7.5 nm between sites 24 and 72; 5.5 nm between sites 24 and 61; and 2 nm between sites 35 and 50. We conclude that alphaS does not retain a "hairpin" structure with two antiparallel helices, as is known to occur with spheroidal micelles, in agreement with our earlier finding that the protein's geometry is determined by the surface topology rather than being constrained by the interhelix linker. While the possibility of local helix discontinuities in the structure of membrane-bound alphaS remains, our data are more consistent with one intact helix. Importantly, we demonstrate that bicelles produce very similar results to liposomes, while offering a major improvement in experimentally accessible distance range and resolution, and thus are an excellent lipid membrane mimetic for the purpose of pulse dipolar ESR spectroscopy.
authors.affiliation: Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Elka R
authors.initials: ER
authors.lastname: Georgieva
authors.forename: Trudy F
authors.initials: TF
authors.lastname: Ramlall
authors.forename: Peter P
authors.initials: PP
authors.lastname: Borbat
authors.forename: Jack H
authors.initials: JH
authors.lastname: Freed
authors.forename: David
authors.initials: D
authors.lastname: Eliezer
mesh_terms: D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D008823:Micelles; D010743:Phospholipids / Q000737:chemistry* / Q000378:metabolism; D017433:Protein Structure, Secondary; D013572:Synaptic Vesicles / Q000737:chemistry / Q000378:metabolism; D053835:Unilamellar Liposomes / Q000737:chemistry; D051844:alpha-Synuclein / Q000737:chemistry* / Q000378:metabolism
chemical_list: D008823:Micelles; D010743:Phospholipids; D053835:Unilamellar Liposomes; D051844:alpha-Synuclein
references.citation: J Biol Chem. 2003 Apr 25;278(17):15313-8
references.pmid: 12586824
references.citation: J Am Chem Soc. 2008 May 28;130(21):6690-1
references.pmid: 18457394
references.citation: Biochemistry. 2007 Apr 17;46(15):4499-509
references.pmid: 17378587
references.citation: Phys Chem Chem Phys. 2007 Apr 28;9(16):1895-910
references.pmid: 17431518
references.citation: J Neurosci. 2005 Nov 23;25(47):10913-21
references.pmid: 16306404
references.citation: Nat Struct Mol Biol. 2006 May;13(5):400-7
references.pmid: 16622408
references.citation: J Mol Biol. 2001 Apr 6;307(4):1061-73
references.pmid: 11286556
references.citation: Biochemistry. 2006 Jul 18;45(28):8453-65
references.pmid: 16834319
references.citation: J Am Chem Soc. 2002 May 15;124(19):5304-14
references.pmid: 11996571
references.citation: PLoS Biol. 2007 Oct;5(10):e271
references.pmid: 17927448
references.citation: Biochemistry. 2005 Jan 11;44(1):329-39
references.pmid: 15628875
references.citation: J Am Chem Soc. 2006 Aug 9;128(31):10004-5
references.pmid: 16881616
references.citation: Protein Sci. 2005 Apr;14(4):862-72
references.pmid: 15741347
references.citation: Biophys J. 2006 Jun 15;90(12):4692-700
references.pmid: 16581836
references.citation: Chem Commun (Camb). 2007 Jan 28;(4):377-9
references.pmid: 17220976
references.citation: Biochemistry. 2004 Apr 27;43(16):4810-8
references.pmid: 15096050
references.citation: Methods Enzymol. 2007;423:52-116
references.pmid: 17609127
references.citation: J Biol Chem. 2005 Mar 11;280(10):9595-603
references.pmid: 15615727
references.citation: J Magn Reson. 2008 Apr;191(2):202-18
references.pmid: 18280189
references.citation: Proc Natl Acad Sci U S A. 2004 Jun 1;101(22):8331-6
references.pmid: 15155902
references.citation: J Neurosci. 1988 Aug;8(8):2804-15
references.pmid: 3411354
references.citation: Q Rev Biophys. 2007 Feb;40(1):1-53
references.pmid: 17565764
references.citation: J Mol Biol. 2003 Jun 13;329(4):763-78
references.pmid: 12787676