title: Dependence of distance distributions derived from double electron-electron resonance pulsed EPR spectroscopy on pulse-sequence time.
abstract: Pulsed double electron-electron resonance (DEER) provides pairwise P(r) distance distributions in doubly spin labeled proteins. We report that in protonated proteins, P(r) is dependent on the length of the second echo period T owing to local environmental effects on the spin-label phase memory relaxation time Tm . For the protein ABD, this effect results in a 1.4 Å increase in the P(r) maximum from T=6 to 20 μs. Protein A has a bimodal P(r) distribution, and the relative height of the shorter distance peak at T=10 μs, the shortest value required to obtain a reliable P(r), is reduced by 40 % relative to that found by extrapolation to T=0. Our results indicate that data at a series of T values are essential for quantitative interpretation of DEER to determine the extent of the T dependence and to extrapolate the results to T=0. Complete deuteration (99 %) of the protein was accompanied by a significant increase in Tm and effectively abolished the P(r) dependence on T.
authors.affiliation: National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520 (USA). jamesbaber@niddk.nih.gov.
authors.forename: James L
authors.initials: JL
authors.lastname: Baber
authors.forename: John M
authors.initials: JM
authors.lastname: Louis
authors.forename: G Marius
authors.initials: GM
authors.lastname: Clore
mesh_terms: D003903:Deuterium / Q000737:chemistry; D041961:Deuterium Exchange Measurement / Q000379:methods*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D008956:Models, Chemical; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D013113:Spin Labels
chemical_list: D011506:Proteins; D013113:Spin Labels; D003903:Deuterium
keywords: EPR spectroscopy; deuteration; distance distributions; phase memory time; proteins