An Improved 15N Relaxation Dispersion Experiment for the Measurement of Millisecond Time-Scale Dynamics in Proteins

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Abstract
A new 15N constant-time relaxation dispersion pulse scheme for the quantification of millisecond time-scale exchange dynamics in proteins is presented. The experiment differs from previously developed sequences in that it includes 1H continuous-wave decoupling during the 15N Carr−Purcell−Meiboom−Gill (CPMG) pulse train that significantly improves the relaxation properties of 15N magnetization, leading to sensitivity gains in experiments. Moreover, it is shown that inclusion of an additional 15N 1800 refocusing pulse (phase cycled ±x) in the center of the CPMG pulse train, consisting of 15Npulses, provides compensation for pulse imperfections beyond the normal CPMG scheme. Relative to existing relaxation-compensated constant-time relaxation dispersion pulse schemes, νCPMG values that are only half as large can be employed, offering increased sensitivity to slow time-scale exchange processes. The robustness of the methodology is illustrated with applications involving a pair of proteins: an SH3 domain that does not show millisecond time-scale exchange and an FF domain with significant chemical exchange contributions.