Rocking motion in solid proteins as studied by solid-state NMR techniques
Biophysics
Experimental and Theoretical Physics of Polymers
Statistical Physics, Nonlinear Dynamics, Complex Systems, Soft and Fluid Matter, Biological Physics
Structural Biology
Final Report Abstract
In order to achieve the goals of the project, we have proposed several methodological improvements to some of the NMR experiments, which allow obtaining precise information on molecular dynamics in the microsecond time scale, free from the interfering effects. We proposed the modification of the CODEX (Centerband-Only Detection of Exchange) pulse sequence, which allows to suppress the influence of the RIDER (Relaxation-Induced Dipolar Exchange with Recoupling) effect. Using this modification, we demonstrated that solid proteins do not undergo slow motion on the millisecond time scale and that the apparent decay of the correlation function observed in this experiment is caused by RIDER. To study faster timescales, we used 5N R1p relaxometry. This method is also prone to an interfering effect such as a spin-spin contribution to the relaxation rate, which reflects spin but not molecular dynamics. To suppress this, we applied proton decoupling during the 15N spin-lock pulse. This modification allowed us to measure the parameters of the slow motion more reliably and accurately than using the conventional relaxation experiment alone. Using this approach, we have shown that the observed slow protein motion has an angular amplitude of only a fewdegrees and a wide distribution of correlation times, ranging from a few to several hundred microseconds. Such slow correlation times are at the very edge of the time window of R1 relaxometry and the obtained parameters of the motion were not very precise. Finally, we applied 2H stimulated echo to measure the correlation function of slow motion, and this experimental approach gave the most reliable, precise and essential results of the whole project. We have proposed a protocol that allows the compensation of distortions in the correlation function obtained with the stimulated echo experiment. We have shown that 15N backbone nuclei and side chain methyl deuterons undergo the same slow motion. In addition to the protein sample, we also performed 2H stimulated echo experiments on microcrystalline tripeptide and an amino acid (L-alanine). Surprisingly, in these samples we observe slow motion in practically the same time range as in the protein sample. In addition, almost no temperature dependence was observed in all three samples. This, together with the independence of the time scale of the motion from the size of the molecules, led us to conclude that this slow motion is in fact ultrasonic phonons, which should be inherent to all rigid biological solids. Thus, the previously used term "rocking" does not reflect the physical nature of this motion, since there is no independent reorientation of molecules within a solid matrix, but rather a correlated bending / twisting of molecular structures resulting in macroscopic elastic vibrations of the whole rigid sample.
Publications
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Relaxation-induced dipolar exchange with recoupling (RIDER) distortions in CODEX experiments. Magnetic Resonance, 1(2), 247-259.
Krushelnitsky, Alexey & Saalwächter, Kay
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Rocking motion in solid proteins studied by the 15N proton-decoupled R1ρ relaxometry. Physical Chemistry Chemical Physics, 25(23), 15885-15896.
Krushelnitsky, Alexey; Hempel, Günter; Jurack, Hannes & Ferreira, Tiago Mendes
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Slow global motions in biosolids studied by the deuteron stimulated echo NMR experiment. The Journal of Chemical Physics, 161(18).
Krushelnitsky, Alexey; Shahsavan, Farhad; Hempel, Günter & Fatkullin, Nail
