ArticleJournal of biomolecular NMR2013
Specific labeling and assignment strategies of valine methyl groups for NMR studies of high molecular weight proteins.
Article in Journal of biomolecular NMR, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 32 papers.
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32 citing papers in PubMed, 64 citations in OpenAlex.
- Extended local deuteration of leucine enhances methyl NMR of large proteins expressed in eukaryotic cells.Chemical communications (Cambridge, England) · 2026Article
- Methyl-specific NMR of therapeutic antibodies: cost-effective isotopic labeling strategies in CHO cells for high-resolution structural characterization.Journal of biomolecular NMR · 2026Article
- Human cells for human proteins: Isotope labeling in mammalian cells in suspension for functional NMR studies.Protein science : a publication of the Protein Society · 2026Article
- A fast and efficient strategy for the NMR assignment of Fab methyl groups.Journal of biomolecular NMR · 2026Article
- A multichaperone condensate enhances protein folding in the endoplasmic reticulum.Nature cell biology · 2025Article
- Leveraging AlphaFold2 and residual dipolar couplings for side-chain methyl group assignment: A case study with S. cerevisiae Xrs2.Journal of magnetic resonance (San Diego, Calif. : 1997) · 2025Article
- Structural basis of substrate recognition and allosteric activation of the proapoptotic mitochondrial HtrA2 protease.Nature communications · 2024Article
- Side-chain dynamics of the αProtein science : a publication of the Protein Society · 2023Article
- Disulfide-Bond-Induced Structural Frustration and Dynamic Disorder in a Peroxiredoxin from MAS NMR.Journal of the American Chemical Society · 2023Article
- Specific isotopic labelling and reverse labelling for protein NMR spectroscopy: using metabolic precursors in sample preparation.Biochemical Society transactions · 2022Review
- Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins.Journal of biomolecular NMR · 2022Article
- Nucleic acid-protein interfaces studied by MAS solid-state NMR spectroscopy.Journal of structural biology: X · 2022Article
- Optimized precursor to simplify assignment transfer between backbone resonances and stereospecifically labelled valine and leucine methyl groups: application to human Hsp90 N-terminal domain.Journal of biomolecular NMR · 2021Article
- Molecular chaperones and their denaturing effect on client proteins.Journal of biomolecular NMR · 2021Article
- Protein Side-Chain-DNA Contacts Probed by Fast Magic-Angle Spinning NMR.The journal of physical chemistry. B · 2020Article
- Spectral editing of intra- and inter-chain methyl-methyl NOEs in protein complexes.Journal of biomolecular NMR · 2020Article
- Aromatic Ring Dynamics, Thermal Activation, and Transient Conformations of a 468 kDa Enzyme by SpecificJournal of the American Chemical Society · 2019Article
- Integrated NMR and cryo-EM atomic-resolution structure determination of a half-megadalton enzyme complex.Nature communications · 2019Article
- Structural investigation of a chaperonin in action reveals how nucleotide binding regulates the functional cycle.Science advances · 2018Article
- Measuring Entropy in Molecular Recognition by Proteins.Annual review of biophysics · 2018Review
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Authors and funding
5 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The specific protonation of valine and leucine methyl groups in proteins is typically achieved by overexpressing proteins in M9/D2O medium supplemented with either labeled α-ketoisovalerate for the labeling of the four prochiral methyl groups or with 2-acetolactate for the stereospecific labeling of the valine and leucine side chains. However, when these labeling schemes are applied to large protein assemblies, significant overlap between the correlations of the valine and leucine methyl groups occurs, hampering the analysis of 2D methyl-TROSY spectra. Analysis of the leucine and valine biosynthesis pathways revealed that the incorporation of labeled precursors in the leucine pathway can be inhibited by the addition of exogenous l-leucine-d10. We exploited this property to label stereospecifically the pro-R and pro-S methyl groups of valine with minimal scrambling to the leucine residues. This new labeling protocol was applied to the 468 kDa homododecameric peptidase TET2 to decrease the complexity of its NMR spectra. All of the pro-S valine methyl resonances of TET2 were assigned by combining mutagenesis with this innovative labeling approach. The assignments were transferred to the pro-R groups using an optimally labeled sample and a set of triple resonance experiments. This improved labeling scheme enables us to overcome the main limitation of overcrowding in the NMR spectra of prochiral methyl groups, which is a prerequisite for the site-specific measurement of the structural and dynamic parameters or for the study of interactions in very large protein assemblies.
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