The spatial effect of protein deuteration on nitroxide spin-label relaxation : Implications for EPR distance measurement
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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.
El Mkami , H , Ward , R J , Bowman , A , Owen-Hughes , T & Norman , D G 2014 , ' The spatial effect of protein deuteration on nitroxide spin-label relaxation : Implications for EPR distance measurement ' Journal of Magnetic Resonance , vol 248 , pp. 36-41 . DOI: 10.1016/j.jmr.2014.09.010
Journal of Magnetic Resonance
Copyright 2014 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/3.0/).
This work was supported by a Wellcome Trust Senior Fellowship (095062) to T.O.-H. The Authors would also like to acknowledge funding from The MRC – United Kingdom, Grant G1100021.
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