Skip to main content
SpringerLink
Log in

13Cα decoupling during direct observation of carbonyl resonances in solution NMR of isotopically enriched proteins

  • Communication
  • Published:
Journal of Biomolecular NMR Aims and scope Submit manuscript

Abstract

Direct detection of 13C can be advantageous when studying uniformly enriched proteins, in particular for paramagnetic proteins or when hydrogen exchange with solvent is fast. A scheme recently introduced for long-observation-window band-selective homonuclear decoupling in solid state NMR, LOW-BASHD (Struppe et al. in J Magn Reson 236:89–94, 2013) is shown to be effective for 13Cα decoupling during direct 13C′ observation in solution NMR experiments too. For this purpose, adjustment of the decoupling pulse parameters and delays is demonstrated to be important for increasing spectral resolution, to reduce three-spin effects, and to decrease the intensity of decoupling side-bands. LOW-BASHD then yields 13C′ line widths comparable to those obtained with the popular IPAP method, while enhancing sensitivity by ca 35 %. As a practical application of LOW-BASHD decoupling, requiring quantitative intensity measurement over a wide dynamic range, the impact of lipid binding on the 13C′-detected NCO spectrum of the intrinsically disordered protein α-synuclein is compared with that on the 1H-detected 1H–15N HSQC spectrum. Results confirm that synuclein’s “dark state” behavior is not caused by paramagnetic relaxation or rapid hydrogen exchange.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  • Barbet-Massin E, Pell AJ, Knight MJ, Webber AL, Felli IC, Pierattelli R, Emsley L, Lesage A, Pintacuda G (2013) 13C-detected through-bond correlation experiments for protein resonance assignment by ultra-fast MAS solid-state NMR. ChemPhysChem 14:3131–3137

    Article  Google Scholar 

  • Barker PB, Shaka AJ, Freeman R (1985) Homonuclear Hartmann–Hahn effects in broad-band decoupling. J Magn Reson 65:535–539

    ADS  Google Scholar 

  • Bax A (1983) Broad-band homonuclear decoupling in heteronuclear shift correlation NMR spectroscopy. J Magn Reson 53:517–520

    ADS  Google Scholar 

  • Bax A, Freeman R (1981) Investigation of complex networks of spin–spin coupling by two-dimensional NMR. J Magn Reson 44:542–561

    ADS  Google Scholar 

  • Bax A, Mehlkopf AF, Smidt J (1979) Homonuclear broad-band decoupled absorption spectra. J Magn Reson 35:167–169

    ADS  Google Scholar 

  • Bermel W, Bertini I, Felli IC, Kummerle R, Pierattelli R (2003) C-13 direct detection experiments on the paramagnetic oxidized monomeric copper, zinc superoxide dismutase. J Am Chem Soc 125:16423–16429

    Article  Google Scholar 

  • Bermel W, Bertini I, Duma L, Felli IC, Emsley L, Pierattelli R, Vasos PR (2005) Complete assignment of heteronuclear protein resonances by protonless NMR spectroscopy. Angew Chem-Int Ed 44:3089–3092

    Article  Google Scholar 

  • Bermel W, Bertini I, Felli IC, Piccioli M, Pierattelli R (2006) C-13-detected protonless NMR spectroscopy of proteins in solution. Prog Nucl Magn Reson Spectrosc 48:25–45

    Article  Google Scholar 

  • Bermel W, Bertini I, Chill J, Felli IC, Haba N, Kumar VMV, Pierattelli R (2012) Exclusively heteronuclear C-13-detected amino-acid-selective NMR experiments for the study of intrinsically disordered proteins (IDPs). ChemBioChem 13:2425–2432

    Article  Google Scholar 

  • Bodner CR, Dobson CM, Bax A (2009) Multiple tight phospholipid-binding modes of alpha-synuclein revealed by solution NMR spectroscopy. J Mol Biol 390:775–790

    Article  Google Scholar 

  • Braunschweiler L, Ernst RR (1983) Coherence transfer by isotropic mixing: application to proton correlation spectroscopy. J Magn Reson 53:521–528

    ADS  Google Scholar 

  • Cavanagh J, Fairbrother WJ, Palmer AG, Rance M, Skelton N (2007) Protein NMR spectroscopy: principles and practice. MA, Elsevier Academic Press, Burlington

    Google Scholar 

  • Delaglio F, Grzesiek S, Vuister GW, Zhu G, Pfeifer J, Bax A (1995) NMRpipe—a multidimensional spectral processing system based on Unix pipes. J Biomol NMR 6:277–293

    Article  Google Scholar 

  • Delsuc MA, Levy GC (1988) The application of maximum-entropy processing to the deconvolution of coupling patterns in NMR. J Magn Reson 76:306–315

    ADS  Google Scholar 

  • Eletsky A, Moreira O, Kovacs H, Pervushin K (2003) A novel strategy for the assignment of side-chain resonances in completely deuterated large proteins using C-13 spectroscopy. J Biomol NMR 26:167–179

    Article  Google Scholar 

  • Eliezer D, Kutluay E, Bussell R, Browne G (2001) Conformational properties of alpha-synuclein in its free and lipid-associated states. J Mol Biol 307:1061–1073

    Article  Google Scholar 

  • Felli IC, Pierattelli R (2014) Novel methods based on C-13 detection to study intrinsically disordered proteins. J Magn Reson 241:115–125

    Article  ADS  Google Scholar 

  • Fusco G, De Simone A, Gopinath T, Vostrikov V, Vendruscolo M, Dobson CM, Veglia G (2014) Direct observation of the three regions in alpha-synuclein that determine its membrane-bound behaviour. Nat commun 5:3827

    Article  ADS  Google Scholar 

  • Garbow JR, Weitekamp DP, Pines A (1982) Bilinear rotation decoupling of homonuclear scalar interactions. Chem Phys Lett 93:504–509

    Article  ADS  Google Scholar 

  • Gil S, Hosek T, Solyom Z, Kuemmerle R, Brutscher B, Pierattelli R, Felli IC (2013) NMR spectroscopic studies of intrinsically disordered proteins at near-physiological conditions. Angewandte Chemie-Int Ed 52:11808–11812

    Article  Google Scholar 

  • Hammarström A, Otting G (1994) Improved spectral resolution in 1H NMR spectroscopy by homonuclear semiselective shaped pulse decoupling during acquisition. J Am Chem Soc 116:8847–8848

    Article  Google Scholar 

  • Levitt MH, Freeman R, Frenkiel T (1982) Broad-band heteronuclear decoupling. J Magn Reson 47:328–330

    ADS  Google Scholar 

  • McCoy MA, Mueller L (1992a) Non-resonant effects of frequency-selective pulses. J Magn Reson 99:18–36

    ADS  Google Scholar 

  • McCoy MA, Mueller L (1992b) Selective shaped pulse decoupling in NMR—homonuclear 13C carbonyl decoupling. J Am Chem Soc 114:2108–2112

    Article  Google Scholar 

  • Meissner A, Duus JO, Sorensen OW (1997) Spin-state-selective excitation. Application for E.COSY-type measurement of J(HH) coupling constants. J Magn Reson 128:92–97

    Article  ADS  Google Scholar 

  • Meyer NH, Zangger K (2013) Simplifying proton NMR spectra by instant homonuclear broadband decoupling. Angew Chem Int Ed 52:7143–7146

    Article  Google Scholar 

  • Nilsson M, Morris GA (2007) Pure shift proton DOSY: diffusion-ordered H-1 spectra without multiplet structure. Chem Commun 9:933–935

    Article  Google Scholar 

  • Sakhaii P, Haase B, Bermel W, Kerssebaum R, Wagner GE, Zangger K (2013) Broadband homodecoupled NMR spectroscopy with enhanced sensitivity. J Magn Reson 233:92–95

    Article  ADS  Google Scholar 

  • Shaka AJ, Barker PB, Freeman R (1987) 3-Spin effects in broad-band decoupling. J Magn Reson 71:520–531

    ADS  Google Scholar 

  • Shaka AJ, Lee CJ, Pines A (1988) Iterative schemes for bilinear operators; application to spin decoupling. J Magn Reson 77:274–293

    ADS  Google Scholar 

  • Struppe JO, Yang C, Wang YC, Hernandez RV, Shamansky LM, Mueller LJ (2013) Long-observation-window band-selective homonuclear decoupling: increased sensitivity and resolution in solid-state NMR spectroscopy of proteins. J Magn Reson 236:89–94

    Article  ADS  Google Scholar 

  • van de Ven FJM, Philippens MEP (1992) Optimization of constant-time evolution in multidimensional NMR experiments. J Magn Reson 97:637–644

    ADS  Google Scholar 

  • Vander Kooi CW, Kupce E, Zuiderweg ERP, Pellecchia M (1999) Line narrowing in spectra of proteins dissolved in a dilute liquid crystalline phase by band-selective adiabatic decoupling: application to H-1(N)-N-15 residual dipolar coupling measurements. J Biomol NMR 15:335–338

    Article  Google Scholar 

  • Vogeli B, Kovacs H, Pervushin K (2005) Simultaneous H-1- or H-2-, N-15- and multiple-band-selective C-13-decoupling during acquisition in C-13-detected experiments with proteins and oligonucleotides. J Biomol NMR 31:1–9

    Article  Google Scholar 

  • Vuister GW, Bax A (1992) Resolution enhancement and spectral editing of uniformly 13C-enriched proteins by homonuclear broadband 13C decoupling. J Magn Reson 98:428–435

    ADS  Google Scholar 

  • Waugh JS (1982) Theory of broad-band spin decoupling. J Magn Reson 50:30–49

    ADS  Google Scholar 

  • Ying J, Roche J, Bax A (2014) Homonuclear decoupling for enhancing resolution and sensitivity in NOE and RDC measurements of peptides and proteins. J Magn Reson 241:97–102

    Article  ADS  Google Scholar 

  • Zangger K, Sterk H (1997) Homonuclear broadband-decoupled NMR spectra. J Magn Reson 124:486–489

    Article  ADS  Google Scholar 

Download references

Acknowledgments

This work was funded by the Intramural Research Program of the National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health (NIH) and the Intramural AIDS-Targeted Antiviral Program of the Office of the Director, NIH. J.H. Lee is the recipient of a KVSTA Fellowship and F. Li acknowledges financial support from the China Scholarship Council.

Author information

Authors and Affiliations

  1. Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA

    Jinfa Ying, Fang Li, Jung Ho Lee & Ad Bax

Authors
  1. Jinfa Ying
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Fang Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jung Ho Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ad Bax
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ad Bax.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 176 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ying, J., Li, F., Lee, J.H. et al. 13Cα decoupling during direct observation of carbonyl resonances in solution NMR of isotopically enriched proteins. J Biomol NMR 60, 15–21 (2014). https://doi.org/10.1007/s10858-014-9853-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10858-014-9853-z

Keywords

Search

Navigation