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Measuring hydrogen exchange in proteins by selective water saturation in 1H–15N SOFAST/BEST-type experiments: advantages and limitations

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Abstract

HETex-SOFAST NMR (Schanda et al. in J Biomol NMR 33:199–211, 2006) has been proposed some years ago as a fast and sensitive method for semi-quantitative measurement of site-specific amide-water hydrogen exchange effects along the backbone of proteins. Here we extend this concept to BEST readout sequences that provide a better resolution at the expense of some loss in sensitivity. We discuss the theoretical background and implementation of the experiment, and demonstrate its performance for an intrinsically disordered protein, 2 well folded globular proteins, and a transiently populated folding intermediate state. We also provide a critical evaluation of the level of accuracy that can be obtained when extracting quantitative exchange rates from HETex NMR measurements.

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References

  • Chevelkov V, Xue Y, Rao DK, Forman-Kay JD, Skrynnikov NR (2010) 15 N H/D-SOLEXSY experiment for accurate measurement of amide solvent exchange rates: application to denatured drkN SH3. J Biomol NMR 46:227–244

    Article  Google Scholar 

  • Croke RL, Sallum CO, Watson E, Watt ED, Alexandrescu AT (2008) Hydrogen exchange of monomeric alpha-synuclein shows unfolded structure persists at physiological temperature and is independent of molecular crowding in Escherichia coli. Protein Sci 17:1434–1445

    Article  Google Scholar 

  • Crowhurst KA, Tollinger M, Forman-Kay JD (2002) Cooperative Interactions and a Non-native Buried Trp in the Unfolded State of an SH3 Domain. J Mol Biol 322:163–178

    Article  Google Scholar 

  • Csizmok V, Felli IC, Tompa P, Banci L, Bertini I (2008) Structural and dynamic characterization of intrinsically disordered human securin by NMR spectroscopy. J Am Chem Soc 130:16873–16879

    Article  Google Scholar 

  • Dempsey CE (2001) Hydrogen exchange in peptides and proteins using NMR spectroscopy. Prog in NMR Spectrosc 39:135–170

    Article  Google Scholar 

  • Englander SW, Kallenbach NR (1983) Hydrogen exchange and structural dynamics of proteins and nucleic acids. Q Rev Biophys 16:521–655

    Article  Google Scholar 

  • Fan J-S, Lim J, Yu B, Yang D (2011) Measurement of amide hydrogen exchange rates with the use of radiation damping. J Biomol NMR 51:151–162

    Article  Google Scholar 

  • Favier A, Brutscher B (2011) Recovering lost magnetization: polarization enhancement in biomolecular NMR. J Biomol NMR 49:9–15

    Article  Google Scholar 

  • Fitzkee NC, Da Torchia, Bax A (2011) Measuring rapid hydrogen exchange in the homodimeric 36 kDa HIV-1 integrase catalytic core domain. Protein Sci 20:500–512

    Article  Google Scholar 

  • Gal M, Schanda P, Brutscher B, Frydman L (2007) UltraSOFAST HMQC NMR and the repetitive acquisition of 2D protein spectra at Hz rates. J Am Chem Soc 129:1372–1377

    Article  Google Scholar 

  • Geen H, Freeman R (1991) Band-selective radiofrequency pulses. J Magn Reson 93:93–141

    ADS  Google Scholar 

  • Gemmecker G, Jahnke W, Kessler H (1993) Measurement of fast proton exchange rates in isotopically labeled compounds. J Am Chem Soc 115:11620–11621

    Article  Google Scholar 

  • Gil S, Hošek T, Solyom Z, Kümmerle R, Brutscher B, Pierattelli R, Felli IC (2013) NMR Spectroscopic Studies of Intrinsically Disordered Proteins at Near-Physiological Conditions. Angew Chem Int Ed Engl 52:11808-11812

  • Grzesiek S, Bax A (1993) Measurement of amide proton-exchange rates and Noes with Water in C-13/N-15-Enriched Calcineurin-B. J Biomol NMR 3:627–638

    Google Scholar 

  • Hwang TL, van Zijl PC, Mori S (1998) Accurate quantitation of water-amide proton exchange rates using the phase-modulated CLEAN chemical EXchange (CLEANEX-PM) approach with a Fast-HSQC (FHSQC) detection scheme. J Biomol NMR 11:221–226

    Article  Google Scholar 

  • Kameda A, Hoshino M, Higurashi T, Takahashi S, Naiki H, Goto Y (2005) Nuclear magnetic resonance characterization of the refolding intermediate of beta(2)-microglobulin trapped by non-native prolyl peptide bond. J Mol Biol 348:383–397

    Article  Google Scholar 

  • Krishnan VV, Murali N (2013) Radiation damping in modern NMR experiments: progress and challenges. Prog in NMR Spectrosc 68:41–57

    Article  Google Scholar 

  • Kupce E, Freeman R (1994) Wide-Band Excitation with Polychromatic Pulses. J Magn Reson A 108:268–273

    Article  ADS  Google Scholar 

  • Kupce E, Boyd J, Campbell ID (1995) Short selective pulses for biochemical applications. J Magn Reson B 106:300–303

    Article  Google Scholar 

  • Lescop E, Schanda P, Brutscher B (2007) A set of BEST triple-resonance experiments for time-optimized protein resonance assignment. J Magn Reson 187:163–169

    Article  ADS  Google Scholar 

  • Mori S, O’Neil Johnson M, Berg JM, Van Zijl PCM (1994) Water exchange filter (WEX Filter) for nuclear magnetic resonance studies of macromolecules. J Am Chem Soc 116:11982–11984

    Article  Google Scholar 

  • Rennella E, Cutuil T, Schanda P, Ayala I, Forge V, Brutscher B (2012) Real-time NMR characterization of structure and dynamics in a transiently populated protein folding intermediate. J Am Chem Soc 134:8066–8069

    Article  Google Scholar 

  • Rennella E et al (2013) Oligomeric states along the folding pathways of β2-microglobulin: kinetics, thermodynamics, and structure. J Mol Biol 425:2722–2736

    Article  Google Scholar 

  • Schanda P, Brutscher B (2005) Very fast two-dimensional NMR spectroscopy for real-time investigation of dynamic events in proteins on the time scale of seconds. J Am Chem Soc 127:8014–8015

    Article  Google Scholar 

  • Schanda P, Kupce E, Brutscher B (2005) SOFAST-HMQC experiments for recording two-dimensional heteronuclear correlation spectra of proteins within a few seconds. J Biomol NMR 33:199–211

    Article  Google Scholar 

  • Schanda P, Forge V, Brutscher B (2006) HET-SOFAST NMR for fast detection of structural compactness and heterogeneity along polypeptide chains. Magn Reson Chem 44:S177–S184

    Article  Google Scholar 

  • Schanda P, Forge V, Brutscher B (2007) Protein folding and unfolding studied at atomic resolution by fast two-dimensional NMR spectroscopy. Proc Natl Acad Sci USA 104:11257–11262

    Article  ADS  Google Scholar 

  • Segawa T, Kateb F, Duma L, Bodenhausen G, Pelupessy P (2008) Exchange rate constants of invisible protons in proteins determined by NMR spectroscopy. ChemBioChem 9:537–542

    Article  Google Scholar 

  • Shishmarev D, Otting G (2011) Radiation damping on cryoprobes. J Magn Reson 213:76–81

    Article  ADS  Google Scholar 

  • Smith AE, Sarkar M, Young GB, Pielak GJ (2013) Amide proton exchange of a dynamic loop in cell extracts. Protein Sci 22:1313–1319

    Google Scholar 

  • Wagner G, Wüthrich K (1982) Amide Proton Exchange and Surface Conformation Basic Pancreatic Trypsin Inhibitor in Solution. J Mol Biol 160:343–361

    Article  Google Scholar 

  • Wider G, Riek R, Wüthrich K (1996) Diffusion filters for separation of solvent—protein and protein–protein nuclear overhauser effects (HYDRA). J Am Chem Soc 7863:11629–11634

    Article  Google Scholar 

  • Yao S, Hinds MG, Murphy JM, Norton RS (2011) Exchange enhanced sensitivity gain for solvent-exchangeable protons in 2D 1H-15 N heteronuclear correlation spectra acquired with band-selective pulses. J Magn Reson 211:243–247

    Article  ADS  Google Scholar 

Download references

Acknowledgments

We thank Isabel Ayala and Karine Giandoreggio for expert protein sample preparation, and we acknowledge support from ANR grant Blanc-InterII-SIMI7-2011 (RNAfolding). This work used the NMR and isotope labeling platforms of the Grenoble Instruct centre (ISBG; UMS 3518 CNRS-CEA-UJF-EMBL) with support from FRISBI (ANR-10-INSB-05-02) and GRAL (ANR-10-LABX-49-01) within the Grenoble Partnership for Structural Biology (PSB).

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Authors and Affiliations

  1. Institut de Biologie Structurale, Université Grenoble 1, 71 avenue des Martyrs, 38044, Grenoble Cedex 9, France

    Enrico Rennella, Zsofia Solyom & Bernhard Brutscher

  2. Commissariat à l’Energie Atomique et aux Energies Alternatives (CEA), Grenoble, France

    Enrico Rennella, Zsofia Solyom & Bernhard Brutscher

  3. Centre National de Recherche Scientifique (CNRS), Grenoble, France

    Enrico Rennella, Zsofia Solyom & Bernhard Brutscher

Authors
  1. Enrico Rennella
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  2. Zsofia Solyom
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  3. Bernhard Brutscher
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Correspondence to Bernhard Brutscher.

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Rennella, E., Solyom, Z. & Brutscher, B. Measuring hydrogen exchange in proteins by selective water saturation in 1H–15N SOFAST/BEST-type experiments: advantages and limitations. J Biomol NMR 60, 99–107 (2014). https://doi.org/10.1007/s10858-014-9857-8

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  • DOI: https://doi.org/10.1007/s10858-014-9857-8

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