Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Fast evaluation of protein dynamics from deficient 15N relaxation data.

ABSTRACT: Simple and convenient method of protein dynamics evaluation from the insufficient experimental 15N relaxation data is presented basing on the ratios, products, and differences of longitudinal and transverse 15N relaxation rates obtained at a single magnetic field. Firstly, the proposed approach allows evaluating overall tumbling correlation time (nanosecond time scale). Next, local parameters of the model-free approach characterizing local mobility of backbone amide N-H vectors on two different time scales, S2 and R ex , can be elucidated. The generalized order parameter, S2, describes motions on the time scale faster than the overall tumbling correlation time (pico- to nanoseconds), while the chemical exchange term, R ex , identifies processes slower than the overall tumbling correlation time (micro- to milliseconds). Advantages and disadvantages of different methods of data handling are thoroughly discussed.

SUBMITTER: Jaremko L

PROVIDER: S-EPMC5953972 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Fast evaluation of protein dynamics from deficient 15N relaxation data.

Jaremko Łukasz Ł Jaremko Mariusz M Ejchart Andrzej A Nowakowski Michał M

Journal of biomolecular NMR 20180328 4

Simple and convenient method of protein dynamics evaluation from the insufficient experimental 15N relaxation data is presented basing on the ratios, products, and differences of longitudinal and transverse 15N relaxation rates obtained at a single magnetic field. Firstly, the proposed approach allows evaluating overall tumbling correlation time (nanosecond time scale). Next, local parameters of the model-free approach characterizing local mobility of backbone amide N-H vec ...[more]

PMID: 29594733

Similar Datasets

15N CPMG Relaxation Dispersion for the Investigation of Protein Conformational Dynamics on the µs-ms Timescale.

Project description:Protein conformational dynamics play fundamental roles in regulation of enzymatic catalysis, ligand binding, allostery, and signaling, which are important biological processes. Understanding how the balance between structure and dynamics governs biological function is a new frontier in modern structural biology and has ignited several technical and methodological developments. Among these, CPMG relaxation dispersion solution NMR methods provide unique, atomic-resolution information on the structure, kinetics, and thermodynamics of protein conformational equilibria occurring on the µs-ms timescale. Here, the study presents detailed protocols for acquisition and analysis of a 15N relaxation dispersion experiment. As an example, the pipeline for the analysis of the µs-ms dynamics in the C-terminal domain of bacteria Enzyme I is shown.

| S-EPMC8168574 | biostudies-literature

Mapping membrane protein backbone dynamics: a comparison of site-directed spin labeling with NMR 15N-relaxation measurements.

Project description:The ability to detect nanosecond backbone dynamics with site-directed spin labeling (SDSL) in soluble proteins has been well established. However, for membrane proteins, the nitroxide appears to have more interactions with the protein surface, potentially hindering the sensitivity to backbone motions. To determine whether membrane protein backbone dynamics could be mapped with SDSL, a nitroxide was introduced at 55 independent sites in a model polytopic membrane protein, TM0026. Electron paramagnetic resonance spectral parameters were compared with NMR (15)N-relaxation data. Sequential scans revealed backbone dynamics with the same trends observed for the R1 relaxation rate, suggesting that nitroxide dynamics remain coupled to the backbone on membrane proteins.

| S-EPMC4190660 | biostudies-literature

Fast-time scale dynamics of outer membrane protein A by extended model-free analysis of NMR relaxation data.

Project description:In order to better understand the dynamics of an integral membrane protein, backbone amide (15)N NMR dynamics measurements of the beta-barrel membrane protein OmpA have been performed at three magnetic fields. A total of nine relaxation data sets were globally analyzed using an extended model-free formalism. The diffusion tensor was found to be prolate axially symmetric with an axial ratio of 5.75, indicating a possible rotation of the protein within the micelle. The generalized order parameters gradually decreased from the mid-plane towards the two ends of the barrel, counteracting the dynamic gradient of the lipids in a matching bilayer, and were dramatically reduced in the extracellular loops. Large-scale internal motions on the ns time scale indicate that entire loops most likely undergo concerted ("sea anemone"-like) motions emanating from their anchoring points on the barrel. The case of OmpA in DPC micelles also illustrates inherent limitations of analyzing the data with even the most sophisticated current models of the model-free formalism. It is likely that conformational exchange processes on the ms-mus also play a role in describing the motions of some residues, but their analysis did not produce unique results that could be independently verified.

| S-EPMC2812607 | biostudies-literature

Measurement of protein backbone 13CO and 15N relaxation dispersion at high resolution.

Project description:Peak overlap in crowded regions of two-dimensional spectra prevents characterization of dynamics for many sites of interest in globular and intrinsically disordered proteins. We present new three-dimensional pulse sequences for measurement of Carr-Purcell-Meiboom-Gill relaxation dispersions at backbone nitrogen and carbonyl positions. To alleviate increase in the measurement time associated with the additional spectral dimension, we use non-uniform sampling in combination with two distinct methods of spectrum reconstruction: compressed sensing and co-processing with multi-dimensional decomposition. The new methodology was validated using disordered protein CD79A from B-cell receptor and an SH3 domain from Abp1p in exchange between its free form and bound to a peptide from the protein Ark1p. We show that, while providing much better resolution, the 3D NUS experiments give the similar accuracy and precision of the dynamic parameters to ones obtained using traditional 2D experiments. Furthermore, we show that jackknife resampling of the spectra yields robust estimates of peak intensities errors, eliminating the need for recording duplicate data points.

| S-EPMC5626786 | biostudies-literature

Water proton spin saturation affects measured protein backbone 15N spin relaxation rates.

Project description:Protein backbone 15N NMR spin relaxation rates are useful in characterizing the protein dynamics and structures. To observe the protein nuclear-spin resonances a pulse sequence has to include a water suppression scheme. There are two commonly employed methods, saturating or dephasing the water spins with pulse field gradients and keeping them unperturbed with flip-back pulses. Here different water suppression methods were incorporated into pulse sequences to measure 15N longitudinal T1 and transversal rotating-frame T1? spin relaxation. Unexpectedly the 15N T1 relaxation time constants varied significantly with the choice of water suppression method. For a 25-kDa Escherichiacoli. glutamine binding protein (GlnBP) the T1 values acquired with the pulse sequence containing a water dephasing gradient are on average 20% longer than the ones obtained using a pulse sequence containing the water flip-back pulse. In contrast the two T1? data sets are correlated without an apparent offset. The average T1 difference was reduced to 12% when the experimental recycle delay was doubled, while the average T1 values from the flip-back measurements were nearly unchanged. Analysis of spectral signal to noise ratios (s/n) showed the apparent slower 15N relaxation obtained with the water dephasing experiment originated from the differences in 1HN recovery for each relaxation time point. This in turn offset signal reduction from 15N relaxation decay. The artifact becomes noticeable when the measured 15N relaxation time constant is comparable to recycle delay, e.g., the 15N T1 of medium to large proteins. The 15N relaxation rates measured with either water suppression schemes yield reasonable fits to the structure. However, data from the saturated scheme results in significantly lower Model-Free order parameters (<S2>=0.81) than the non-saturated ones (<S2>=0.88), indicating such order parameters may be previously underestimated.

| S-EPMC3339441 | biostudies-literature

Impact of two-bond 15N-15N scalar couplings on 15N transverse relaxation measurements for arginine side chains of proteins.

Project description:NMR relaxation of arginine (Arg) 15Nε nuclei is useful for studying side-chain dynamics of proteins. In this work, we studied the impact of two geminal 15N-15N scalar couplings on measurements of transverse relaxation rates (R 2 ) for Arg side-chain 15Nε nuclei. For 12 Arg side chains of the DNA-binding domain of the Antp protein, we measured the geminal 15N-15N couplings ( 2 J NN ) of the 15Nε nuclei and found that the magnitudes of the 2 J NN coupling constants were virtually uniform with an average of 1.2 Hz. Our simulations, assuming ideal 180° rotations for all 15N nuclei, suggested that the two 2 J NN couplings of this magnitude could in principle cause significant modulation in signal intensities during the Carr-Purcell-Meiboom-Gill (CPMG) scheme for Arg 15Nε R 2 measurements. However, our experimental data show that the expected modulation via two 2 J NN couplings vanishes during the 15N CPMG scheme. This quenching of J modulation can be explained by the mechanism described in Dittmer and Bodenhausen (Chemphyschem 7:831-836, 2006). This effect allows for accurate measurements of R 2 relaxation rates for Arg side-chain 15Nε nuclei despite the presence of two 2 J NN couplings. Although the so-called recoupling conditions may cause overestimate of R 2 rates for very mobile Arg side chains, such conditions can readily be avoided through appropriate experimental settings.

| S-EPMC6020141 | biostudies-literature

Characterization of the fast dynamics of protein amino acid side chains using NMR relaxation in solution.

Project description: Not available

| S-EPMC2547146 | biostudies-literature

Conformational dynamics of recoverin's Ca2+-myristoyl switch probed by 15N NMR relaxation dispersion and chemical shift analysis.

Project description:Recoverin, a member of the neuronal calcium sensor (NCS) branch of the calmodulin superfamily, serves as a calcium sensor in retinal rod cells. Ca(2+) -induced conformational changes in recoverin promote extrusion of its covalently attached myristate, known as the Ca(2+)-myristoyl switch. Here, we present nuclear magnetic resonance (NMR) relaxation dispersion and chemical shift analysis on (15) N-labeled recoverin to probe main chain conformational dynamics. (15) N NMR relaxation data suggest that Ca(2+)-free recoverin undergoes millisecond conformational dynamics at particular amide sites throughout the protein. The addition of trace Ca(2+) levels (0.05 equivalents) increases the number of residues that show detectable relaxation dispersion. The Ca(2+)-dependent chemical shifts and relaxation dispersion suggest that recoverin has an intermediate conformational state (I) between the sequestered apo state (T) and Ca(2+) saturated extruded state (R): T ↔ I ↔ R. The first step is a fast conformational equilibrium ([T]/[I] < 100) on the millisecond time scale (τ(ex) δω < 1). The final step (I ↔ R) is much slower (τ(ex) δω > 1). The main chain structure of I is similar in part to the structure of half-saturated E85Q recoverin with a sequestered myristoyl group. We propose that millisecond dynamics during T ↔ I may transiently increase the exposure of Ca(2+)-binding sites to initiate Ca(2+) binding that drives extrusion of the myristoyl group during I ↔ R.

| S-EPMC3092842 | biostudies-literature

Overall rotational diffusion and internal mobility in domain II of protein G from Streptococcus determined from 15N relaxation data.

Project description:The backbone dynamics and overall tumbling of protein G have been investigated using 15N relaxation. Comparison of measured R2/R1 relaxation rate ratios with known three-dimensional coordinates of the protein show that the rotational diffusion tensor is significantly asymmetric, exhibiting a prolate axial symmetry. Extensive Monte Carlo simulations have been used to estimate the uncertainty due to experimental error in the relaxation rates to be D(parallel)/D(perpendicular) = 1.68 +/- 0.08, while the dispersion in the NMR ensemble leads to a variation of D(parallel)/D(perpendicular) = 1.65 +/- 0.03. Incorporation of this tensorial description into a Lipari-Szabo type analysis of internal motion has allowed us to accurately describe the local dynamics of the molecule. This analysis differs from an earlier study where the overall rotational diffusion was described by a spherical top. In this previous analysis, exchange parameters were fitted to many of the residues in the alpha helix. This was interpreted as reflecting a small motion of the alpha helix with respect to the beta sheet. We propose that the differential relaxation properties of this helix compared to the beta sheet are due to the near-orthogonality of the NH vectors in the two structural motifs with respect to the unique axis of the diffusion tensor. Our analysis shows that when anisotropic rotational diffusion is taken into account NH vectors in these structural motifs appear to be equally rigid. This study underlines the importance of a correct description of the rotational diffusion tensor if internal motion is to be accurately investigated.

| S-EPMC2144651 | biostudies-other

Variability of the 15N chemical shielding tensors in the B3 domain of protein G from 15N relaxation measurements at several fields. Implications for backbone order parameters.

Project description:We applied a combination of 15N relaxation and CSA/dipolar cross-correlation measurements at five magnetic fields (9.4, 11.7, 14.1, 16.4, and 18.8 T) to determine the 15N chemical shielding tensors for backbone amides in protein G in solution. The data were analyzed using various model-independent approaches and those based on Lipari-Szabo approximation, all of them yielding similar results. The results indicate a range of site-specific values of the anisotropy (CSA) and orientation of the 15N chemical shielding tensor, similar to those in ubiquitin (Fushman, et al. J. Am. Chem. Soc. 1998, 120, 10947; J. Am. Chem. Soc. 1999, 121, 8577). Assuming a Gaussian distribution of the 15N CSA values, the mean anisotropy is -173.9 to -177.2 ppm (for 1.02 A NH bond length) and the site-to-site CSA variability is +/-17.6 to +/-21.4 ppm, depending on the method used. This CSA variability is significantly larger than derived previously for ribonuclease H (Kroenke, et al. J. Am. Chem. Soc. 1999, 121, 10119) or recently, using "meta-analysis" for ubiquitin (Damberg, et al. J. Am. Chem. Soc. 2005, 127, 1995). Standard interpretation of 15N relaxation studies of backbone dynamics in proteins involves an a priori assumption of a uniform 15N CSA. We show that this assumption leads to a significant discrepancy between the order parameters obtained at different fields. Using the site-specific CSAs obtained from our study removes this discrepancy and allows simultaneous fit of relaxation data at all five fields to Lipari-Szabo spectral densities. These findings emphasize the necessity of taking into account the variability of 15N CSA for accurate analysis of protein dynamics from 15N relaxation measurements.

| S-EPMC2519110 | biostudies-literature