Solid State NMR Relaxation Studies of Triosephosphate Isomerase
By: Caitlin Quinn
Publisher: [publisher not identified]
Published: 2013
Place of publication: [New York, N.Y.?]
Language: eng
Description: Both protein structure and dynamics are essential to understanding biological function. NMR is a powerful technique for the observation of protein dynamics in that dynamics can be observed site-specifically over a wide range of timescales from picoseconds to seconds. Spin relaxation measurements, including relaxation in the rotating frame (R1ρ), can be very sensitive to exchange processes in proteins, particularly on the millisecond-to-microsecond timescale. Using solid state NMR, few techniques exist that can quantify dynamics on this timescale. Previous R1ρ relaxation measurements in the solid state have utilized reorientation of a dipole tensor to observe dynamics. This application is limited to systems where the nucleus of interest has an attached proton. Relaxation studies using the reorientation of a chemical shift tensor are applicable to a broader range of systems. Furthermore, solid state experiments do not require a change in the isotropic chemical shift as is necessary in solution NMR.

We combined R1ρ measurements of the model compound dimethyl sulfone (DMS) with data-fitting routines in Spinevolution to show that R1ρ relaxation due to reorientation of a chemical shift tensor is a large effect in the solid state and these measurements can be used to quantify chemical exchange processes. The temperature dependence of the exchange rates determined with R1ρ measurements is in agreement with other measurements of the dynamics of DMS with various solid state NMR techniques. Deuteration and sparse isotropic labeling were necessary to obtain quantitative results. To distinguish the exchange contribution to relaxation from other effects (R2 relaxation), low temperatures and high spin-lock field strengths were utilized. R1ρ experiments and magic angle spinning (MAS) one-dimensional spectra were used to characterize phosphate ligand binding in the glycolytic protein triosephosphate isomerase. 1D spectra indicated the presence of both isotropic and anisotropic phosphate populations.

These states included an unbound state with an isotropic chemical shift tensor, and a protein-bound state in which the anisotropic features are reintroduced through chelation with protein backbone amides. The chemical shift anisotropy tensor of the bound phosphate ligand was fit using spinning sideband analysis of slow MAS spectra and suggest the ligand is in a dianionic state. The temperature dependence of R1ρ measurements indicated a fast dynamic process above the microsecond timescale at physiological temperatures.
Notes: Department: Chemistry.

Thesis advisor: Ann McDermott.

Thesis (Ph.D.)--Columbia University, 2013.