NMR Spectroscopy for Monitoring Functional Dynamics in Solution


Protein functional dynamics are defined as the atomic thermal fluctuations or the segmental motions that are essential for the function of a biomolecule. NMR is a very versatile technique that is well suited for obtaining quantitative information from these processes at atomic resolution and in multiple timescales. This article focuses on recent NMR developments to study functional dynamics, making special incidence in the experiments aimed for the characterisation of chemical–conformational exchange in the microsecond to millisecond timescale. In a second section, the novel‐solution NMR techniques that combine chemical exchange with saturation transfer experiments are addressed to finish with an overview on the dynamic information that can be extracted from residual dipolar couplings.

Key Concepts

  • NMR is well suited to study biomolecular dynamics at multiple timescales.
  • Many enzymatic reactions occur in the microsecond to millisecond timescale.
  • Relaxation dispersion experiments can extract quantitative information about the exchange process.
  • CEST and DEST experiments provide useful information on conformations that are invisible in the NMR spectrum.
  • Residual dipolar couplings contain information about spin dynamics that can be extracted and quantified.

Keywords: protein functional dynamics; NMR spectroscopy; residual dipolar couplings; CEST; DEST; conformational exchange; relaxation dispersion experiments; spin‐lock field

Figure 1. (a) Spin relaxation experiment cover most of the timescales where the effective protein internal (and overall) motions take place. Faster librations can be detected by nuclear spin relaxation measurements (R1, R2 and NOE) or by analysing the dynamic contribution of the residual dipolar couplings (RDCs). The line‐shape analysis and the relaxation dispersion experiments account for motions involving a larger number of atoms and are most relevant in biology. The slower motions can be characterised in NMR using EXSY and ZZ exchange experiments. (b) Owing to their intrinsic flexibility, proteins sample a range of thermodynamically accessible conformations within a hierarchy of timescales. The population of each state is determined by its Gibbs free energy, while the inter‐conversion between states is owed to the energy barriers that separate them. Three examples of the continuum of timescales are depicted in the figure for clarity: fast bond librations occurring in the picosecond (ps) to nanosecond (ns) timescale, segmental motions involving a larger number of atoms occurring in the microsecond (µs) to millisecond (ms) timescale and large domain rearrangements, occurring in slow timescales (s).
Figure 2. CPMG relaxation dispersion experiments for three different residues of the bacterial gene repressor YmoA. The transversal relaxation rate is measured as a function of the pulse train frequency. The solid lines correspond to the best fitting to the relaxation dispersion profile. Different residues show different sensitivities to the experiment and show no dispersion (red), moderate exchange contribution (blue) and a large contribution (black) of chemical exchange to line broadening.
Figure 3. Residual dipolar couplings. (a) Schematic representation of protein (red blob) in anisotropic alignment medium (vertical cylinders) with respect to the magnetic field B0. (b) Dynamics probed by RDC, the Azz, Ayy and Axx axes represents the alignment tensor for nuclear dipole pairs (bond vector with white and black balls) in a protein depicted in blue and the relative orientation of same dipole pairs in different alignment condition, where the protein is shown as red and golden yellow. The different accessible space available for the dipole pairs makes it feasible to obtain dynamics by RDC.


Ban D, Sabo TM, Griesinger C and Lee D (2013) Molecules 18: 11904.

Barnett JA (1998) Yeast 14: 1439.

Bertini I, Kursula P, Luchinat C, et al. (2009) Journal of the American Chemical Society 131: 5134.

Bhabha G, Lee J, Ekiert DC, et al. (2011) Science 332: 234.

Boehr DD, McElheny D, Dyson HJ and Wright PE (2006) Science 313: 1638.

Bouvignies G, Vallurupalli P, Hansen DF, et al. (2011) Nature 477: 111.

Bouvignies G and Kay LE (2012a) Journal of Biomolecular NMR 53: 303.

Bouvignies G and Kay LE (2012b) Journal of Physical Chemistry B 116: 14311.

Fawzi NL, Ying J, Ghirlando R, Torchia DA and Clore GM (2011) Nature 480: 268.

Fawzi NL, Ying J, Torchia DA and Clore GM (2012) Nature Protocols 7: 1523.

Hansen DF, Neudecker P, Vallurupalli P, Mulder FA and Kay LE (2010) Journal of the American Chemical Society 132: 42.

Kleckner IR and Foster MP (2011) Biochimica et Biophysica Acta 1814: 942.

Korzhnev DM, Vernon RM, Religa TL, et al. (2011) Journal of the American Chemical Society 133: 10974.

Lange OF, Lakomek NA, Fares C, et al. (2008) Science 320: 1471.

Lee J‐S, Parasoglou P, Xia D, Jerschow A and Regatte RR (2013) Scientific Reports: 3.

Louie A (2013) Journal of Magnetic Resonance Imaging 38: 530.

McConnell HM (1958) The Journal of Chemical Physics 28: 430.

Millet O, Loria JP, Kroenke CD, Pons M and Palmer AG (2000) Journal of the American Chemical Society 122: 2867.

Mittermaier AK and Kay LE (2009) Trends in Biochemical Sciences 34: 601.

Mulder FA, Skrynnikov NR, Hon B, Dahlquist FW and Kay LE (2001) Journal of the American Chemical Society 123: 967.

Palmer AG 3rd (2014) Journal of Magnetic Resonance 241: 3.

Sekhar A and Kay LE (2013) Proceedings of the National Academy of Sciences of the United States of America 110: 12867.

Tolman JR, Flanagan JM, Kennedy MA and Prestegard JH (1997) Nature Structural Biology 4: 292.

Tolman JR and Ruan K (2006) Chemical Reviews 106: 1720.

Trott O and Palmer AG (2002) Journal of Magnetic Resonance 154: 157.

Ulmer TS, Ramirez BE, Delaglio F and Bax A (2003) Journal of the American Chemical Society 125: 9179.

Vallurupalli P, Bouvignies G and Kay LE (2012) Journal of the American Chemical Society 134: 8148.

van Zijl PCM and Yadav NN (2011) Magnetic Resonance in Medicine 65: 927.

Further Reading

Mittermaier A and Kay LE (2006) Science 312: 224.

Palmer AG 3rd (2004) Chem Rev 104: 3623.

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Sivanandam, VN, and Millet, Oscar(Apr 2015) NMR Spectroscopy for Monitoring Functional Dynamics in Solution. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0003104.pub2]