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

Deuterium (H) spin relaxation of CHD methyl groups has been widely applied to investigate picosecond-to-nanosecond conformational dynamics in proteins by solution-state NMR spectroscopy. The B dependence of the H spin relaxation rates is represented by a linear relationship between the spectral density function at three discrete frequencies J(0), J(ω) and J(2ω). In this study, the linear relation between H relaxation rates at B 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 C-C labeled sample of E. coli RNase H. Deuterium relaxation rate constants (R, R, R, R) were measured for 57 well-resolved CHD moieties in RNase H at H frequencies of 475 MHz, 500 MHz, 900 MHz, and 950 MHz. The spectral density mapping of the 475/950 MHz data combination was performed independently and jointly to validate the expected relationship between data recorded at B fields separated by a factor of two. The final analysis was performed by jointly analyzing 475/950 MHz rates with 700 MHz rates interpolated from 500/900 MHz data to yield six J(ω) values for each methyl peak. The J(ω) profile for each peak was fit to the original (τ, S, τ) or extended model-free function (τ, S, S, τ, τ) to obtain optimized dynamic parameters.