title: The spatial effect of protein deuteration on nitroxide spin-label relaxation: implications for EPR distance measurement.
abstract: Pulsed electron-electron double resonance (PELDOR) coupled with site-directed spin labeling is a powerful technique for the elucidation of protein or nucleic acid, macromolecular structure and interactions. The intrinsic high sensitivity of electron paramagnetic resonance enables measurement on small quantities of bio-macromolecules, however short relaxation times impose a limit on the sensitivity and size of distances that can be measured using this technique. The persistence of the electron spin-echo, in the PELDOR experiment, is one of the most crucial limitations to distance measurement. At a temperature of around 50 K one of the predominant factors affecting persistence of an echo, and as such, the sensitivity and measurable distance between spin labels, is the electron spin echo dephasing time (Tm). It has become normal practice to use deuterated solvents to extend Tm and recently it has been demonstrated that deuteration of the underlying protein significantly extends Tm. Here we examine the spatial effect of segmental deuteration of the underlying protein, and also explore the concentration and temperature dependence of highly deuterated systems.
authors.affiliation: School of Physics and Astronomy, University of St. Andrews, St. Andrews KY16 9SS, UK.
authors.forename: Hassane
authors.initials: H
authors.lastname: El Mkami
authors.affiliation: Nucleic Acids Structure Research Group, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Richard
authors.initials: R
authors.lastname: Ward
authors.affiliation: Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Andrew
authors.initials: A
authors.lastname: Bowman
authors.affiliation: Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Tom
authors.initials: T
authors.lastname: Owen-Hughes
authors.affiliation: Nucleic Acids Structure Research Group, University of Dundee, Dundee DD1 5EH, UK. Electronic address: d.g.norman@dundee.ac.uk.
authors.forename: David G
authors.initials: DG
authors.lastname: Norman
mesh_terms: D000595:Amino Acid Sequence; D003903:Deuterium / Q000032:analysis / Q000737:chemistry*; D041961:Deuterium Exchange Measurement / Q000379:methods*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D006657:Histones / Q000737:chemistry* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D009589:Nitrogen Oxides / Q000032:analysis* / Q000737:chemistry*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D062211:Spatio-Temporal Analysis; D013113:Spin Labels
chemical_list: D006657:Histones; D009589:Nitrogen Oxides; D013113:Spin Labels; D003903:Deuterium
keywords: DEER; Deuteration; EPR; PELDOR; Relaxation; Spin-label; T(m)