Analysis of local molecular motions of aromatic sidechains in proteins by 2D and 3D fast MAS NMR spectroscopy and quantum mechanical calculations

Paluch, Piotr; Pawlak, T.; Jeziorna, Agata; Trébosc, Julien; Hou, Guangjin; Vega, Alexander J.; Amoureux, Jean‐Paul; Dračínský, Martin; Polenova, Tatyana; Potrzebowski, Marek J.

doi:10.1039/c5cp04475h

Cited by 20 publications

(55 citation statements)

References 56 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…1 H- 13 C dipolar parameters can be conveniently measured in a 3D experiment, where the dipolar recoupling period is followed by a 13 C- 13 C mixing step, such as fpRFDR, as reported previously 32 and shown in Figure 1c in the context of wPARS recoupling sequence.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Improving dipolar recoupling for site-specific structural and dynamics studies in biosolids NMR: windowed RN-symmetry sequences

Zhang

et al. 2016

Phys. Chem. Chem. Phys.

Self Cite

View full text Add to dashboard Cite

Experimental characterization of one-bond heteronuclear dipolar couplings is essential for structural and dynamics characterization of molecules by solid-state NMR. Accurate measurement of heteronuclear dipolar tensor parameters in magic angle spinning NMR requires that the recoupling sequences efficiently reintroduce the desired heteronuclear dipolar coupling term, fully suppress other interactions (such as chemical shift anisotropy and homonuclear dipolar couplings), and be insensitive to experimental imperfections, such as radio frequency (rf) field mismatch. In this study, we demonstrate that the introduction of window delays into the basic elements of a PARS sequence results in a greatly improved protocol, termed windowed PARS (wPARS), which yields clean dipolar lineshapes that are unaffected by other spin interactions and are largely insensitive to experimental imperfections. Higher dipolar scaling factor can be attained in this technique with respect to PARS, which is particularly useful for the measurement of relatively small dipolar couplings. The advantages of wPARS are verified experimentally on model molecules N-Acetyl-Valine (NAV) and a tripeptide Met-Leu-Phe (MLF). The incorporation of wPARS into 3D heteronuclear correlation experiments permits accurate site-specific determination of dipolar tensors in proteins, as demonstrated on dynein light chain 8 (LC8). Through 3D wPARS recoupling based spectroscopy we have determined both backbone and sidechain dipolar tensors in LC8 in a residue-resolved manner. We discuss these in the context of conformational dynamics of LC8. We have addressed the effect of paramagnetic relaxant Cu(II)-EDTA doping on the dipolar coupling parameters in LC8 and observed no significant differences with respect to the neat sample permitting fast data collection. Our results indicate that wPARS is advantageous with respect to the windowless version of the sequence and is applicable to a broad range of systems including but not limited to biomolecules.

show abstract

Section: Resultsmentioning

confidence: 99%

“…This result indicates that the fpRFDR mixing does not introduce any artifacts into the spectra, in accord with the previous findings for a 3D CPVC-RFDR experiment. 32 …”

Section: Resultsmentioning

confidence: 99%

Improving dipolar recoupling for site-specific structural and dynamics studies in biosolids NMR: windowed RN-symmetry sequences

Zhang

et al. 2016

Phys. Chem. Chem. Phys.

Self Cite

View full text Add to dashboard Cite

show abstract

“…The evidence of such dynamics under non-DNP conditions has been recently reported for aromatic sidechains. 63-67 In principle, analysis of the conformational space and the populations could provide a glimpse into the type of motions that exist. However, data at variable temperatures will be required for a quantitative analysis of such dynamics, which can depend on motional timescales, amplitudes, populations and potential energy barriers.…”

Section: Resultsmentioning

confidence: 99%

Dynamic Nuclear Polarization Enhanced MAS NMR Spectroscopy for Structural Analysis of HIV-1 Protein Assemblies

Gupta

Hou

et al. 2016

J. Phys. Chem. B

Self Cite

View full text Add to dashboard Cite

Mature infectious HIV-1 virions contain conical capsids comprised of CA protein, generated by the proteolytic cleavage cascade of the Gag polyprotein, termed maturation. The mechanism of capsid core formation through the maturation process remains poorly understood. We present DNP-enhanced MAS NMR studies of tubular assemblies of CA and Gag CA-SP1 maturation intermediate and report 20 – 64 fold sensitivity enhancements due to DNP at 14.1 T. These sensitivity enhancements enabled direct observation of spacer peptide 1 (SP1) resonances in CA-SP1 by dipolar based correlation experiments, unequivocally indicating that the SP1 peptide is unstructured in assembled CA-SP1 at cryogenic temperatures, corroborating our earlier results. Furthermore, the dependence of DNP enhancements and spectral resolution on magnetic field strength (9.4 – 18.8 T) and temperature (109 – 180 K) was investigated. Our results suggest that DNP-based measurements could potentially provide residue-specific dynamics information by allowing for the extraction of temperature dependence of the anisotropic tensorial or relaxation parameters. With DNP, we were able to detect multiple well-resolved isoleucine sidechain conformers, unique intermolecular correlations across two CA molecules, and functionally relevant conformationally disordered states such as the 14-residue SP1 peptide, none of which are visible at ambient temperatures. The detection of isolated conformers and intermolecular correlations can provide crucial constraints for structure determination of these assemblies. Overall, our results establish DNP-based MAS NMR as an excellent tool for characterization of HIV-1 assemblies.

show abstract

“…109,110,111,112,113,114 Its fast MAS variant allows to add 1 H-13 C dipolar dimensions in multidimensional experiments 68,115,116,117,118 Recently, it was demonstrated that on-resonance CW 14 N irradiation during the CSA recoupling period with rf field ~30 times smaller than the 14 N quadrupolar couplings largely decouples 14 N-1 H interactions leading to perfect 1 H CSA lineshapes 127 .…”

Section: Sensitivitymentioning

confidence: 99%

Fast magic-angle sample spinning solid-state NMR at 60–100 kHz for natural abundance samples

Nishiyama

2016

Solid State Nuclear Magnetic Resonance

130

239

View full text Add to dashboard Cite

Analysis of local molecular motions of aromatic sidechains in proteins by 2D and 3D fast MAS NMR spectroscopy and quantum mechanical calculations

Cited by 20 publications

References 56 publications

Improving dipolar recoupling for site-specific structural and dynamics studies in biosolids NMR: windowed RN-symmetry sequences

Improving dipolar recoupling for site-specific structural and dynamics studies in biosolids NMR: windowed RN-symmetry sequences

Dynamic Nuclear Polarization Enhanced MAS NMR Spectroscopy for Structural Analysis of HIV-1 Protein Assemblies

Fast magic-angle sample spinning solid-state NMR at 60–100 kHz for natural abundance samples

Contact Info

Product

Resources

About