@article{20258,
  abstract     = {The specific introduction of ^1H-^13C or ^1H-^15N moieties into otherwise deuterated proteins holds great potential for high-resolution solution and magic-angle spinning (MAS) NMR studies of protein structure and dynamics. Arginine residues play key roles for example at active sites of enzymes. Taking advantage of a chemically synthesized Arg with a ^13C-^1H2 group in an otherwise deuterated backbone, we demonstrate here the usefulness of proton-detected MAS NMR approaches to probe arginine dynamics. In experiments with crystalline ubiquitin and the 134 kDa tetrameric enzyme malate dehydrogenase we detected a wide range of motions, from sites that are rigid on time scales of at least tens of milliseconds to residues undergoing predominantly nanosecond motions. Spin-relaxation and dipolar-coupling measurements enabled quantitative determination of these dynamics. We observed microsecond dynamics of residue Arg54 in crystalline ubiquitin, whose backbone is known to sample different β-turn conformations on this time scale. The labeling scheme and experiments presented here expand the toolkit for high-resolution proton-detected MAS NMR.},
  author       = {Rohden, Darja and Napoli, Federico and Kapitonova, Anna and Tatman, Benjamin and Lichtenecker, Roman J. and Schanda, Paul},
  issn         = {1089-8638},
  journal      = {Journal of Molecular Biology},
  number       = {23},
  publisher    = {Elsevier},
  title        = {{Arginine dynamics probed by magic-angle spinning NMR with a specific isotope-labeling scheme}},
  doi          = {10.1016/j.jmb.2025.169379},
  volume       = {437},
  year         = {2025},
}

@article{20538,
  abstract     = {In this study, we describe an integrated approach for methyl group assignment comprising precursor-based selective methyl group labeling, a novel pulse sequence for methyl to backbone coherence transfer and chemical shift predictions using UCBShift 2.0. The utility of this novel α-ketoacid isotopologue is shown by the adaptation of an HMBC-HMQC pulse sequence that simultaneously connects geminal methyl groups of leucine and valine residues to each other and to the protein backbone. By additional 13C,2H-labeling of residues other than valine and leucine residues of the protein, important chemical shift information about neighboring residues (following valine and leucine residues) can be achieved. Thus, different valine and leucine residues in a protein can be characterized as a specific chemical shift vector. Frequency matching with predicted chemical shifts via UCBShift 2.0 using experimental data taken from a subset of the BMRB database revealed a correct assignment performance of about 90%. With applications to proteins of 60.2 kDa and 134 kDa (4 × 33.5 kDa) in size, we demonstrate that the approach provides valuable information even for very large proteins.},
  author       = {Knödlstorfer, Sonja and Toscano, Giorgia and Ptaszek, Aleksandra L. and Kontaxis, Georg and Napoli, Federico and Schneider, Jakob and Maier, Katharina and Kapitonova, Anna and Lichtenecker, Roman J. and Schanda, Paul and Konrat, Robert},
  issn         = {1089-8638},
  journal      = {Journal of Molecular Biology},
  number       = {23},
  publisher    = {Elsevier},
  title        = {{A novel HMBC-CC-HMQC NMR strategy for methyl assignment using triple-13C-labeled α-ketoisovalerate integrated with UCBShift 2.0}},
  doi          = {10.1016/j.jmb.2025.169465},
  volume       = {437},
  year         = {2025},
}

@article{15401,
  abstract     = {Amide-proton-detected magic-angle-spinning NMR of deuterated proteins has become a main technique in NMR-based structural biology. In standard deuteration protocols that rely on D2O-based culture media, non-exchangeable amide sites remain deuterated, making these sites unobservable. Here we demonstrate that proteins produced with a H2O-based culture medium doped with deuterated cell lysate allow scientists to overcome this “reprotonation bottleneck” while retaining a high level of deuteration (ca. 80 %) and narrow linewidths. We quantified coherence lifetimes of several proteins prepared with this labeling pattern over a range of magic-angle-spinning (MAS) frequencies (40–100 kHz). We demonstrate that under commonly used conditions (50–60 kHz MAS), the amide 1H linewidths with our labeling approach are comparable to those of perdeuterated proteins and better than those of protonated samples at 100 kHz. For three proteins in the 33–50 kDa size range, many previously unobserved amides become visible. We report how to prepare the deuterated cell lysate for our approach from fractions of perdeuterated cultures which are usually discarded, and we show that such media can be used identically to commercial media. The residual protonation of Hα sites allows for well-resolved Hα-detected spectra and Hα resonance assignment, exemplified by the de novo assignment of 168 Hα sites in a 39 kDa protein. The approach based on this H2O/cell-lysate deuteration and MAS frequencies compatible with 1.3 or 1.9 mm rotors presents a strong sensitivity benefit over 0.7 mm 100 kHz MAS experiments.},
  author       = {Napoli, Federico and Guan, Jia-Ying and Arnaud, Charles-Adrien and Macek, Pavel and Fraga, Hugo and Breyton, Cécile and Schanda, Paul},
  issn         = {2699-0016},
  journal      = {Magnetic Resonance},
  number       = {1},
  pages        = {33--49},
  publisher    = {Copernicus Publications},
  title        = {{Deuteration of proteins boosted by cell lysates: High-resolution amide and Ha magic-angle-spinning (MAS) NMR without the reprotonation bottleneck}},
  doi          = {10.5194/mr-5-33-2024},
  volume       = {5},
  year         = {2024},
}

@article{14036,
  abstract     = {Magic-angle spinning (MAS) nuclear magnetic resonance (NMR) is establishing itself as a powerful method for the characterization of protein dynamics at the atomic scale. We discuss here how R1ρ MAS relaxation dispersion NMR can explore microsecond-to-millisecond motions. Progress in instrumentation, isotope labeling, and pulse sequence design has paved the way for quantitative analyses of even rare structural fluctuations. In addition to isotropic chemical-shift fluctuations exploited in solution-state NMR relaxation dispersion experiments, MAS NMR has a wider arsenal of observables, allowing to see motions even if the exchanging states do not differ in their chemical shifts. We demonstrate the potential of the technique for probing motions in challenging large enzymes, membrane proteins, and protein assemblies.},
  author       = {Napoli, Federico and Becker, Lea Marie and Schanda, Paul},
  issn         = {1879-033X},
  journal      = {Current Opinion in Structural Biology},
  number       = {10},
  publisher    = {Elsevier},
  title        = {{Protein dynamics detected by magic-angle spinning relaxation dispersion NMR}},
  doi          = {10.1016/j.sbi.2023.102660},
  volume       = {82},
  year         = {2023},
}