title: Time-resolved EPR immersion depth studies of a transmembrane peptide incorporated into bicelles.
abstract: The reduction in EPR signal intensity of nitroxide spin-labels by ascorbic acid has been measured as a function of time to investigate the immersion depth of the spin-labeled M2δ AChR peptide incorporated into a bicelle system utilizing EPR spectroscopy. The corresponding decay curves of n-DSA (n=5, 7, 12, and 16) EPR signals have been used to (1) calibrate the depth of the bicelle membrane and (2) establish a calibration curve for measuring the depth of spin-labeled transmembrane peptides. The kinetic EPR data of CLS, n-DSA (n=5, 7, 12, and 16), and M2δ AChR peptide spin-labeled at Glu-1 and Ala-12 revealed excellent exponential and linear fits. For a model M2δ AChR peptide, the depth of immersion was calculated to be 5.8Å and 3Å for Glu-1, and 21.7Å and 19Å for Ala-12 in the gel-phase (298K) and L(α)-phases (318K), respectively. The immersion depth values are consistent with the pitch of an α-helix and the structural model of M2δ AChR incorporated into the bicelle system is in a good agreement with previous studies. Therefore, this EPR time-resolved kinetic technique provides a new reliable method to determine the immersion depth of membrane-bound peptides, as well as, explore the structural characteristics of the M2δ AChR peptide.
authors.affiliation: Walsh University, North Canton, OH 44720, USA.
authors.forename: Nisreen A
authors.initials: NA
authors.lastname: Nusair
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Mayo
authors.forename: Tia D
authors.initials: TD
authors.lastname: Dorozenski
authors.forename: Thomas B
authors.initials: TB
authors.lastname: Cardon
authors.forename: Johnson J
authors.initials: JJ
authors.lastname: Inbaraj
authors.forename: Ethan S
authors.initials: ES
authors.lastname: Karp
authors.forename: Justin P
authors.initials: JP
authors.lastname: Newstadt
authors.forename: Stuart M
authors.initials: SM
authors.lastname: Grosser
authors.forename: Gary A
authors.initials: GA
authors.lastname: Lorigan
mesh_terms: D000818:Animals; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D006801:Humans; D008081:Liposomes / Q000737:chemistry*; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry*; D017433:Protein Structure, Secondary; D043585:Receptor, Muscarinic M2 / Q000737:chemistry*; D013113:Spin Labels
chemical_list: D008081:Liposomes; D010455:Peptides; D043585:Receptor, Muscarinic M2; D013113:Spin Labels
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