title: Probing the structure of membrane proteins with electron spin echo envelope modulation spectroscopy.
abstract: A new approach has been developed to probe the structural properties of membrane peptides and proteins using the pulsed electron paramagnetic resonance technique of electron spin echo envelope modulation (ESEEM) spectroscopy and the α-helical M2δ subunit of the acetylcholine receptor incorporated into phospholipid bicelles. To demonstrate the practicality of this method, a cysteine-mutated nitroxide spin label (SL) is positioned 1, 2, 3, and 4 residues away from a fully deuterated Val side chain (denoted i + 1 to i + 4). The characteristic periodicity of the α-helical structure gives rise to a unique pattern in the ESEEM spectra. In the i + 1 and i + 2 samples, the ²H nuclei are too far away to be detected. However, with the 3.6 residue per turn pattern of an α-helix, the i + 3 and i + 4 samples reveal a strong signal from the ²H nuclei of the Val side chain. Modeling studies verify these data suggesting that the closest ²H-labeled Val to SL distance would in fact be expected in the i + 3 and i + 4 samples. This technique is very advantageous, because it provides pertinent qualitative structural information on an inherently difficult system like membrane proteins in a short period of time (minutes) with small amounts of protein (μg).
authors.affiliation: Department of Chemistry and Biochemistry, Miami University, Oxford, Ohio, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Mayo
authors.forename: Andy
authors.initials: A
authors.lastname: Zhou
authors.forename: Indra
authors.initials: I
authors.lastname: Sahu
authors.forename: Robert
authors.initials: R
authors.lastname: McCarrick
authors.forename: Parker
authors.initials: P
authors.lastname: Walton
authors.forename: Adam
authors.initials: A
authors.lastname: Ring
authors.forename: Kaylee
authors.initials: K
authors.lastname: Troxel
authors.forename: Aaron
authors.initials: A
authors.lastname: Coey
authors.forename: Jaclyn
authors.initials: J
authors.lastname: Hawn
authors.forename: Abdul-Hamid
authors.initials: AH
authors.lastname: Emwas
authors.forename: Gary A
authors.initials: GA
authors.lastname: Lorigan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D029941:Fish Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D043585:Receptor, Muscarinic M2 / Q000737:chemistry*; D013113:Spin Labels; D014101:Torpedo / Q000378:metabolism*
chemical_list: D029941:Fish Proteins; D043585:Receptor, Muscarinic M2; D013113:Spin Labels