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Protein NMR pp 353-373 | Cite as

Covariance NMR Processing and Analysis for Protein Assignment

  • Bradley J. Harden
  • Dominique P. Frueh
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1688)

Abstract

During NMR resonance assignment it is often necessary to relate nuclei to one another indirectly, through their common correlations to other nuclei. Covariance NMR has emerged as a powerful technique to correlate such nuclei without relying on error-prone peak peaking. However, false-positive artifacts in covariance spectra have impeded a general application to proteins. We recently introduced pre- and postprocessing steps to reduce the prevalence of artifacts in covariance spectra, allowing for the calculation of a variety of 4D covariance maps obtained from diverse combinations of pairs of 3D spectra, and we have employed them to assign backbone and sidechain resonances in two large and challenging proteins. In this chapter, we present a detailed protocol describing how to (1) properly prepare existing 3D spectra for covariance, (2) understand and apply our processing script, and (3) navigate and interpret the resulting 4D spectra. We also provide solutions to a number of errors that may occur when using our script, and we offer practical advice when assigning difficult signals. We believe such 4D spectra, and covariance NMR in general, can play an integral role in the assignment of NMR signals.

Key words

NMR Covariance Resonance assignment Peak lists Spectral derivative 4D spectra 

Notes

Acknowledgments

The Frueh lab is supported by the National Institute of Health, grant R01GM104257.

References

  1. 1.
    Sprangers R, Kay LE (2007) Quantitative dynamics and binding studies of the 20S proteasome by NMR. Nature 445:618–622. doi: 10.1038/nature05512 CrossRefPubMedGoogle Scholar
  2. 2.
    Tugarinov V, Kay LE (2003) Ile, Leu, and Val methyl assignments of the 723-residue malate synthase G using a new labeling strategy and novel NMR methods. J Am Chem Soc 125:13868–13878. doi: 10.1021/ja030345s CrossRefPubMedGoogle Scholar
  3. 3.
    Brüschweiler R, Zhang F (2004) Covariance nuclear magnetic resonance spectroscopy. J Chem Phys 120:5253–5260. doi: 10.1063/1.1647054 CrossRefPubMedGoogle Scholar
  4. 4.
    Brüschweiler R (2004) Theory of covariance nuclear magnetic resonance spectroscopy. J Chem Phys 121:409–414. doi: 10.1063/1.1755652 CrossRefPubMedGoogle Scholar
  5. 5.
    Trbovic N, Smirnov S, Zhang F, Brüschweiler R (2004) Covariance NMR spectroscopy by singular value decomposition. J Magn Reson 171:277–283. doi: 10.1016/j.jmr.2004.08.007 CrossRefPubMedGoogle Scholar
  6. 6.
    Zhang F, Brüschweiler R (2004) Spectral deconvolution of chemical mixtures by covariance NMR. ChemPhysChem 5:794–796. doi: 10.1002/cphc.200301073 CrossRefPubMedGoogle Scholar
  7. 7.
    Zhang F, Brüschweiler R (2004) Indirect covariance NMR spectroscopy. J Am Chem Soc 126:13180–13181. doi: 10.1021/ja047241h CrossRefPubMedGoogle Scholar
  8. 8.
    Blinov KA, Larin NI, Kvasha MP et al (2005) Analysis and elimination of artifacts in indirect covariance NMR spectra via unsymmetrical processing. Magn Reson Chem 43:999–1007. doi: 10.1002/mrc.1674 CrossRefPubMedGoogle Scholar
  9. 9.
    Blinov KA, Larin NI, Williams AJ et al (2006) Long-range carbon-carbon connectivity via unsymmetrical indirect covariance processing of HSQC and HMBC NMR data. Magn Reson Chem 44:107–109. doi: 10.1002/mrc.1766 CrossRefPubMedGoogle Scholar
  10. 10.
    Kupče E, Freeman R (2006) Hyperdimensional NMR spectroscopy. J Am Chem Soc 128:6020–6021. doi: 10.1021/ja0609598 CrossRefPubMedGoogle Scholar
  11. 11.
    Lescop E, Brutscher B (2007) Hyperdimensional protein NMR spectroscopy in peptide-sequence space. J Am Chem Soc 129:11916–11917. doi: 10.1021/ja0751577 CrossRefPubMedGoogle Scholar
  12. 12.
    Benison G, Berkholz DS, Barbar E (2007) Protein assignments without peak lists using higher-order spectra. J Magn Reson 189:173–181. doi: 10.1016/j.jmr.2007.09.009 CrossRefPubMedGoogle Scholar
  13. 13.
    Chen K, Delaglio F, Tjandra N (2010) A practical implementation of cross-spectrum in protein backbone resonance assignment. J Magn Reson 203:208–212. doi: 10.1016/j.jmr.2009.12.018 CrossRefPubMedGoogle Scholar
  14. 14.
    Snyder DA, Brüschweiler R (2009) Generalized indirect covariance NMR formalism for establishment of multidimensional spin correlations. J Phys Chem A 113:12898–12903. doi: 10.1021/jp9070168 CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Snyder DA, Ghosh A, Zhang F et al (2008) Z-matrix formalism for quantitative noise assessment of covariance nuclear magnetic resonance spectra. J Chem Phys 129:1–9. doi: 10.1063/1.2975206 CrossRefGoogle Scholar
  16. 16.
    Snyder DA, Zhang F, Brüschweiler R (2007) Covariance NMR in higher dimensions: application to 4D NOESY spectroscopy of proteins. J Biomol NMR 39:165–175. doi: 10.1007/s10858-007-9187-1 CrossRefPubMedGoogle Scholar
  17. 17.
    Snyder DA, Xu Y, Yang D, Brüschweiler R (2007) Resolution-enhanced 4D 15N/13C NOESY protein NMR spectroscopy by application of the covariance transform. J Am Chem Soc 129:14126–14127. doi: 10.1021/ja075533n CrossRefPubMedGoogle Scholar
  18. 18.
    Harden B, Nichols S, Frueh D (2014) Facilitated assignment of large protein NMR signals with covariance sequential spectra using spectral derivatives. J Am Chem Soc 136:13106–13109CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Harden BJ, Mishra SH, Frueh DP (2015) Effortless assignment with 4D covariance sequential correlation maps. J Magn Reson 260:83–88. doi: 10.1016/j.jmr.2015.09.007 CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Mishra SH, Frueh D (2015) Assignment of methyl NMR resonances of a 52 kDa protein with residue-specific 4D correlation maps. J Biomol NMR 62:281–290. doi: 10.1007/s10858-015-9943-6 CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Keller R (2004) The computer aided resonance assignment tutorial. Cantina Verlag, Goldau (Switzerland), p 225Google Scholar
  22. 22.
    Short T, Alzapiedi L, Brüschweiler R, Snyder D (2011) A covariance NMR toolbox for MATLAB and OCTAVE. J Magn Reson 209:75–78. doi: 10.1016/j.jmr.2010.11.018 CrossRefPubMedGoogle Scholar
  23. 23.
    The MathWorks Inc. (2014) MATLABGoogle Scholar
  24. 24.
    Eaton JW, Bateman D, Hauberg S, Wehbring R (2015) GNU Octave version 4.0.0 manual: a high-level interactive language for numerical computations. ISBN 1441413006, URL http://www.gnu.org/software/octave/doc/interpreter/

Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  1. 1.Department of Biophysics and Biophysical ChemistryJohns Hopkins University School of MedicineBaltimoreUSA

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