Probing Transient Conformational States of Proteins by Solid‐State R_1ρ Relaxation‐Dispersion NMR Spectroscopy

Abstract

The function of proteins depends on their ability to sample a variety of states differing in structure and free energy. Deciphering how the various thermally accessible conformations are connected, and understanding their structures and relative energies is crucial in rationalizing protein function. Many biomolecular reactions take place within microseconds to milliseconds, and this timescale is therefore of central functional importance. Here we show that R1ρ relaxation dispersion experiments in magic-angle-spinning solid-state NMR spectroscopy make it possible to investigate the thermodynamics and kinetics of such exchange process, and gain insight into structural features of short-lived states.

Studying dynamics by magic-angle spinning solid-state NMR spectroscopy: Principles and applications to biomolecules

“…Prior to the study of Lewandowski et al mentioned above, there have also been reports of measurements of 15 N R 1ϱ relaxation-rate constants in highly-deuterated proteins at lower MAS frequency of about 10-20 kHz [113], and over the last years several other studies of R 1ϱ rate constants have been reported under different conditions of sample deuteration, MAS frequency, and RF-field strengths [45,100,110,133,134]. The conditions under which coherent dephasing is sufficiently suppressed depends on the choice of the labeling scheme (in particular deuteration), the MAS frequency, and the RF field strength.…”

“…In the case where the two exchanging states differ also in the orientations of dipolarcoupling and CSA tensors, the situation becomes more complex: as the RF-field amplitude approaches the rotary-resonance conditions [139,140] (ν 1 = n ν r where n = 1,2), we observe an increased R 1ϱ decay-rate constant [45,133,134]. This is expected, as the spectral density at the difference between MAS frequency and the RF-field amplitude is relevant for R 1ϱ (see Eq.…”

“…It is similar to the MAS dependence of the crosscorrelated relaxation rate constants discussed above (see Figure 17 and Figure 18). The dependence of the R 1ϱ rate constant on MAS frequency and RF field strength has been exploited previously to study microsecond motion [45,133,137].…”

“…The expected evolution of R 1ϱ with the RF-field amplitude is thus the sum of the two limiting cases, a Bloch-McConnell-type of dispersion, and a dispersion in the vicinity of the rotary-resonance conditions. Solid-state R 1ϱ relaxation-dispersion experiments have been shown to probe isotropic chemical-shift changes and bond-vector changes in proteins [45].…”

Structure of fully protonated proteins by proton-detected magic-angle spinning NMR

“…This opens the way to rapid sequential assignment of backbone resonances (24)(25)(26)(27), as well as to the unambiguous measurement of detailed structural and dynamical parameters (28)(29)(30)(31)(32). A further increase in the MAS frequency to 100 kHz allows resonance assignment (20), a structure determination of a model protein (16), and interaction studies (15) with as little as 0.5 mg of sample.…”