Browse
Submit Data
Databases
API
Help

Dataset Information

13 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Optimized selection of slow-relaxing ¹³C transitions in methyl groups of proteins: application to relaxation dispersion.

ABSTRACT: Optimized selection of the slow-relaxing components of single-quantum ¹³C magnetization in ¹³CH₃ methyl groups of proteins using acute (< 90°) angle ¹H radio-frequency pulses, is described. The optimal selection scheme is more relaxation-tolerant and provides sensitivity gains in comparison to the experiment where the undesired (fast-relaxing) components of ¹³C magnetization are simply 'filtered-out' and only 90° ¹H pulses are employed for magnetization transfer to and from ¹³C nuclei. When applied to methyl ¹³C single-quantum Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion experiments for studies of chemical exchange, the selection of the slow-relaxing ¹³C transitions results in a significant decrease in intrinsic (exchange-free) transverse spin relaxation rates of all exchanging species. For exchanging systems involving high-molecular-weight species, the lower transverse relaxation rates translate into an increase in the information content of the resulting relaxation dispersion profiles.

SUBMITTER: Tugarinov V

PROVIDER: S-EPMC7704780 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Accounting for the temperature dependence of 13C spin-lattice relaxation of methyl groups in the glycyl-alanyl-leucine model system under MAS with spin diffusion.

Project description:The difficulties in quantitatively modeling the temperature dependence of spin-lattice relaxation in a model isotope-enriched peptide are explored as a prelude to obtaining dynamics parameters for motions in proteins from such measurements. The degree to which this can be handled by adding spin diffusion to a bath in standard rate matrix relaxation theory is studied using a small tri-peptide model system, glycyl-alanyl-leucine (GAL). We observe in this molecule that the relaxation of backbone carbons CO and C? is not dominated by local fluctuations of the 13C-1H dipolar couplings, but rather by 13C-13C spin diffusion to nearby methyl relaxation sinks. A treatment of the methyl relaxation itself, which ignores 13C-13C spin diffusion effects back to the otherwise slowly relaxing bath, provides poor agreement between theory and experimental data obtained for the temperature dependence of the methyl relaxation rates. Closed form approximate spectral densities and relaxation rates for a methyl group during magic angle spinning are obtained to compute the needed transition rates. These average computed rates, in conjunction with an extended form of the Solomon equations, are found to adequately model the temperature dependence of the methyl relaxation rates when spin diffusion is included. The barrier to rotation for the alanine methyl in GAL is determined to be 3.5 kcal mol-1.

| S-EPMC6817761 | biostudies-literature

Quantifying the effects of long-range 13C-13C dipolar coupling on measured relaxation rates in RNA.

Project description:Selective stable isotope labeling has transformed structural and dynamics analysis of RNA by NMR spectroscopy. These methods can remove 13C-13C dipolar couplings that complicate 13C relaxation analyses. While these phenomena are well documented for sites with adjacent 13C nuclei (e.g. ribose C1'), less is known about so-called isolated sites (e.g. adenosine C2). To investigate and quantify the effects of long-range (> 2 Å) 13C-13C dipolar interactions on RNA dynamics, we simulated adenosine C2 relaxation rates in uniformly [U-13C/15N]-ATP or selectively [2-13C]-ATP labeled RNAs. Our simulations predict non-negligible 13C-13C dipolar contributions from adenosine C4, C5, and C6 to C2 longitudinal (R1) relaxation rates in [U-13C/15N]-ATP labeled RNAs. Moreover, these contributions increase at higher magnetic fields and molecular weights to introduce discrepancies that exceed 50%. This will become increasingly important at GHz fields. Experimental R1 measurements in the 61 nucleotide human hepatitis B virus encapsidation signal ε RNA labeled with [U-13C/15N]-ATP or [2-13C]-ATP corroborate these simulations. Thus, in the absence of selectively labeled samples, long-range 13C-13C dipolar contributions must be explicitly taken into account when interpreting adenosine C2 R1 rates in terms of motional models for large RNAs.

| S-EPMC8131303 | biostudies-literature

Measurement of Ligand-Target Residence Times by 1H Relaxation Dispersion NMR Spectroscopy.

Project description:A ligand-observed 1H NMR relaxation experiment is introduced for measuring the binding kinetics of low-molecular-weight compounds to their biomolecular targets. We show that this approach, which does not require any isotope labeling, is applicable to ligand-target systems involving proteins and nucleic acids of variable molecular size. The experiment is particularly useful for the systematic investigation of low affinity molecules with residence times in the micro- to millisecond time regime.

| S-EPMC5150660 | biostudies-literature

A suite of 19F based relaxation dispersion experiments to assess biomolecular motions.

Project description:Proteins and nucleic acids are highly dynamic bio-molecules that can populate a variety of conformational states. NMR relaxation dispersion (RD) methods are uniquely suited to quantify the associated kinetic and thermodynamic parameters. Here, we present a consistent suite of 19F-based CPMG, on-resonance R1ρ and off-resonance R1ρ RD experiments. We validate these experiments by studying the unfolding transition of a 7.5 kDa cold shock protein. Furthermore we show that the 19F RD experiments are applicable to very large molecular machines by quantifying dynamics in the 360 kDa half-proteasome. Our approach significantly extends the timescale of chemical exchange that can be studied with 19F RD, adds robustness to the extraction of exchange parameters and can determine the absolute chemical shifts of excited states. Importantly, due to the simplicity of 19F NMR spectra, it is possible to record complete datasets within hours on samples that are of very low costs. This makes the presented experiments ideally suited to complement static structural information from cryo-EM and X-ray crystallography with insights into functionally relevant motions.

| S-EPMC7701166 | biostudies-literature

Slow ring flips in aromatic cluster of GB1 studied by aromatic 13C relaxation dispersion methods.

Project description:Ring flips of phenylalanine and tyrosine are a hallmark of protein dynamics. They report on transient breathing motions of proteins. In addition, flip rates also depend on stabilizing interactions in the ground state, like aromatic stacking or cation-? interaction. So far, experimental studies of ring flips have almost exclusively been performed on aromatic rings without stabilizing interactions. Here we investigate ring flip dynamics of Phe and Tyr in the aromatic cluster in GB1. We found that all four residues of the cluster, Y3, F30, Y45 and F52, display slow ring flips. Interestingly, F52, the central residue of the cluster, which makes aromatic contacts with all three others, is flipping significantly faster, while the other rings are flipping with the same rates within margin of error. Determined activation enthalpies and activation volumes of these processes are in the same range of other reported ring flips of single aromatic rings. There is no correlation of the number of aromatic stacking interactions to the activation enthalpy, and no correlation of the ring's extent of burying to the activation volume. Because of these findings, we speculate that F52 is undergoing concerted ring flips with each of the other rings.

| S-EPMC7080667 | biostudies-literature

Conformational exchange of aromatic side chains characterized by L-optimized TROSY-selected ¹³C CPMG relaxation dispersion.

Project description:Protein dynamics on the millisecond time scale commonly reflect conformational transitions between distinct functional states. NMR relaxation dispersion experiments have provided important insights into biologically relevant dynamics with site-specific resolution, primarily targeting the protein backbone and methyl-bearing side chains. Aromatic side chains represent attractive probes of protein dynamics because they are over-represented in protein binding interfaces, play critical roles in enzyme catalysis, and form an important part of the core. Here we introduce a method to characterize millisecond conformational exchange of aromatic side chains in selectively (13)C labeled proteins by means of longitudinal- and transverse-relaxation optimized CPMG relaxation dispersion. By monitoring (13)C relaxation in a spin-state selective manner, significant sensitivity enhancement can be achieved in terms of both signal intensity and the relative exchange contribution to transverse relaxation. Further signal enhancement results from optimizing the longitudinal relaxation recovery of the covalently attached (1)H spins. We validated the L-TROSY-CPMG experiment by measuring fast folding-unfolding kinetics of the small protein CspB under native conditions. The determined unfolding rate matches perfectly with previous results from stopped-flow kinetics. The CPMG-derived chemical shift differences between the folded and unfolded states are in excellent agreement with those obtained by urea-dependent chemical shift analysis. The present method enables characterization of conformational exchange involving aromatic side chains and should serve as a valuable complement to methods developed for other types of protein side chains.

| S-EPMC3427480 | biostudies-literature

Correction to: A suite of 19F based relaxation dispersion experiments to assess biomolecular motions.

Project description:In the original publication, Figures 3 and 6 were displayed incorrectly due to a mistake made by the publisher. The correct version of Figs. 3 and 6 are given below.

| S-EPMC7852850 | biostudies-literature

Probing conformational transitions towards mutagenic Watson-Crick-like G·T mismatches using off-resonance sugar carbon R1? relaxation dispersion.

Project description:NMR off-resonance R1? relaxation dispersion measurements on base carbon and nitrogen nuclei have revealed that wobble G·T/U mismatches in DNA and RNA duplexes exist in dynamic equilibrium with short-lived, low-abundance, and mutagenic Watson-Crick-like conformations. As Watson-Crick-like G·T mismatches have base pairing geometries similar to Watson-Crick base pairs, we hypothesized that they would mimic Watson-Crick base pairs with respect to the sugar-backbone conformation as well. Using off-resonance R1? measurements targeting the sugar C3' and C4' nuclei, a structure survey, and molecular dynamics simulations, we show that wobble G·T mismatches adopt sugar-backbone conformations that deviate from the canonical Watson-Crick conformation and that transitions toward tautomeric and anionic Watson-Crick-like G·T mismatches restore the canonical Watson-Crick sugar-backbone. These measurements also reveal kinetic isotope effects for tautomerization in D2O versus H2O, which provide experimental evidence in support of a transition state involving proton transfer. The results provide additional evidence in support of mutagenic Watson-Crick-like G·T mismatches, help rule out alternative inverted wobble conformations in the case of anionic G·T-, and also establish sugar carbons as new non-exchangeable probes of this exchange process.

| S-EPMC7508749 | biostudies-literature

Determining methyl sidechain conformations in a CS-ROSETTA model using methyl 1H-13C residual dipolar couplings.

Project description:Modelling of protein structures based on backbone chemical shifts, using programs such as CS-ROSETTA, is becoming increasingly popular, especially for systems where few restraints are available or where homologous structures are already known. While the reliability of CS-ROSETTA calculations can be improved by incorporation of some additional backbone NMR data such as those afforded by residual dipolar couplings or minimal NOE data sets involving backbone amide protons, the sidechain conformations are largely modelled by statistical energy terms. Here, we present a simple method based on methyl residual dipolar couplings that can be used to determine the rotameric state of the threefold symmetry axis of methyl groups that occupy a single rotamer, determine rotameric distributions, and identify regions of high flexibility. The method is demonstrated for methyl side chains of a deletion variant of the human chaperone DNAJB6b.

| S-EPMC7083688 | biostudies-literature

Ferrotoroidic ground state in a heterometallic {CrIIIDyIII6} complex displaying slow magnetic relaxation.

Project description:Toroidal quantum states are most promising for building quantum computing and information storage devices, as they are insensitive to homogeneous magnetic fields, but interact with charge and spin currents, allowing this moment to be manipulated purely by electrical means. Coupling molecular toroids into larger toroidal moments via ferrotoroidic interactions can be pivotal not only to enhance ground state toroidicity, but also to develop materials displaying ferrotoroidic ordered phases, which sustain linear magneto-electric coupling and multiferroic behavior. However, engineering ferrotoroidic coupling is known to be a challenging task. Here we have isolated a {CrIIIDyIII6} complex that exhibits the much sought-after ferrotoroidic ground state with an enhanced toroidal moment, solely arising from intramolecular dipolar interactions. Moreover, a theoretical analysis of the observed sub-Kelvin zero-field hysteretic spin dynamics of {CrIIIDyIII6} reveals the pivotal role played by ferrotoroidic states in slowing down the magnetic relaxation, in spite of large calculated single-ion quantum tunneling rates.

| S-EPMC5647347 | biostudies-literature