Deuterium spin relaxation of fractionally deuterated ribonuclease H using paired 475 and 950Â*MHz NMR spectrometers

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Deuterium spin relaxation of fractionally deuterated ribonuclease H using paired 475 and 950Â*MHz NMR spectrometers

Abstract

Deuterium (2H) spin relaxation of 13CH2D methyl groups has been widely applied to investigate picosecond-to-nanosecond conformational dynamics in proteins by solution-state NMR spectroscopy. The B0 dependence of the 2H spin relaxation rates is represented by a linear relationship between the spectral density function at three discrete frequencies J(0), J(Ï?D) and J(2Ï?D). In this study, the linear relation between 2H relaxation rates at B0 fields separated by a factor of two and the interpolation of rates at intermediate frequencies are combined for a more robust approach for spectral density mapping. The general usefulness of the approach is demonstrated on a fractionally deuterated (55%) and alternate 13C-12C labeled sample of E. coli RNase H. Deuterium relaxation rate constants (R1, R1Ï?, RQ, RAP) were measured for 57 well-resolved 13CH2D moieties in RNase H at 1H frequencies of 475Â*MHz, 500Â*MHz, 900Â*MHz, and 950Â*MHz. The spectral density mapping of the 475/950Â*MHz data combination was performed independently and jointly to validate the expected relationship between data recorded at B0 fields separated by a factor of two. The final analysis was performed by jointly analyzing 475/950Â*MHz rates with 700Â*MHz rates interpolated from 500/900Â*MHz data to yield six J(Ï?D) values for each methyl peak. The J(Ï?) profile for each peak was fit to the original (Ï?M, Sf2, Ï?f) or extended model-free function (Ï?M, Sf2, Ss2, Ï?f, Ï?s) to obtain optimized dynamic parameters.



Source: Journal of Biomolecular NMR