15N transverse relaxation measurements for the characterization of µs–ms dynamics are deteriorated by the deuterium isotope effect on 15N resulting from solvent exchange
- Journal Article
Rights / licenseIn Copyright - Non-Commercial Use Permitted
¹⁵N R₂ relaxation measurements are key for the elucidation of the dynamics of both folded and intrinsically disordered proteins (IDPs). Here we show, on the example of the intrinsically disordered protein α-synuclein and the folded domain PDZ2, that at physiological pH and near physiological temperatures amide—water exchange can severely skew Hahn-echo based ¹⁵N R₂ relaxation measurements as well as low frequency data points in CPMG relaxation dispersion experiments. The nature thereof is the solvent exchange with deuterium in the sample buffer, which modulates the ¹⁵N chemical shift tensor via the deuterium isotope effect, adding to the apparent relaxation decay which leads to systematic errors in the relaxation data. This results in an artificial increase of the measured apparent ¹⁵N R₂ rate constants—which should not be mistaken with protein inherent chemical exchange contributions, Rex, to ¹⁵N R₂. For measurements of ¹⁵N R₂ rate constants of IDPs and folded proteins at physiological temperatures and pH, we recommend therefore the use of a very low D₂O molar fraction in the sample buffer, as low as 1%, or the use of an external D₂O reference along with a modified ¹⁵N R₂ Hahn-echo based experiment. This combination allows for the measurement of Rₑₓ contributions to ¹⁵N R₂ originating from conformational exchange in a time window from µs to ms. Show more
Journal / seriesJournal of Biomolecular NMR
Pages / Article No.
SubjectIntrinsically disordered proteins; NMR relaxation experiments; Amide exchange; Deuterium isotope effect; Loop dynamics
Organisational unit03782 - Riek, Roland / Riek, Roland
03496 - Meier, Beat H. (emeritus) / Meier, Beat H. (emeritus)
NotesIt was possible to publish this article open access thanks to a Swiss National Licence with the publisher.
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