Enhanced sensitivity for pulse dipolar EPR spectroscopy using variable-time RIDME
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Pulse dipolar spectroscopy (PDS) measurements are an important complementary tool in structural biology and are increasingly applied to macromolecular assemblies implicated in human health and disease at physiological concentrations. This requires ever higher sensitivity, and recent advances have driven PDS measurements into the mid-nanomolar concentration regime, though optimization and acquisition of such measurements remains experimentally demanding and time expensive. One important consideration is that constant-time acquisition represents a hard limit for measurement sensitivity, depending on the maximum measured distance. Determining this distance a priori has been facilitated by machine-learning structure prediction (AlphaFold2 and RoseTTAFold) but is often confounded by non-representative behaviour in frozen solution that may mandate multiple rounds of optimization and acquisition. Herein, we endeavour to simultaneously enhance sensitivity and streamline PDS measurement optimization to one-step by benchmarking a variable-time acquisition RIDME experiment applied to CuII-nitroxide and CuII-CuII model systems. Results demonstrate marked sensitivity improvements of both 5- and 6-pulse variable-time RIDME of between 2- and 5-fold over the constant-time analogues.
Wort , J , Ackermann , K , Giannoulis , A & Bode , B E 2023 , ' Enhanced sensitivity for pulse dipolar EPR spectroscopy using variable-time RIDME ' , Journal of Magnetic Resonance , vol. 352 , 107460 . https://doi.org/10.1016/j.jmr.2023.107460
Journal of Magnetic Resonance
Copyright 2023 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY license.
DescriptionFunding: For the purpose of open access, the authors have applied a Creative Commons Attribution (CC BY) licence to any Accepted Author Manuscript version arising. The authors thank the StAnD (St Andrews and Dundee) EPR group for long-standing support and in particular Dr El Mkami for assistance with PDS experiments. J. L.W. acknowledges support by the BBSRC DTP Eastbio (BB/M010996/1). A. G. acknowledges the EPSRC-funded Centre for Doctoral Training in ‘integrated magnetic resonance’, iMR-CDT (EP/J500045/1) B. E. B. and K. A. acknowledge support by the Leverhulme Trust (RPG-2018-397). B. E. B. acknowledges equipment funding by BBSRC (BB/R013780/1 and BB/T017740/1).
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