Journal of Biomolecular NMR

, Volume 52, Issue 1, pp 57–64 | Cite as

Amide temperature coefficients in the protein G B1 domain

  • Jennifer H. Tomlinson
  • Mike P. WilliamsonEmail author


Temperature coefficients have been measured for backbone amide 1H and 15N nuclei in the B1 domain of protein G (GB1), using temperatures in the range 283–313 K, and pH values from 2.0 to 9.0. Many nuclei display pH-dependent coefficients, which were fitted to one or two pKa values. 1H coefficients showed the expected behaviour, in that hydrogen-bonded amides have less negative values, but for those amides involved in strong hydrogen bonds in regular secondary structure there is a negative correlation between strength of hydrogen bond and size of temperature coefficient. The best correlation to temperature coefficient is with secondary shift, indicative of a very approximately uniform thermal expansion. The largest pH-dependent changes in coefficient are for amides in loops adjacent to sidechain hydrogen bonds rather than the amides involved directly in hydrogen bonds, indicating that the biggest determinant of the temperature coefficient is temperature-dependent loss of structure, not hydrogen bonding. Amide 15N coefficients have no clear relationship with structure.


Temperature coefficient Chemical shift Hydrogen bond pH Protein G 



We thank the Biotechnology and Biological Science Research Council (UK) for funding a studentship to JHT.


  1. Alexander P, Fahnestock S, Lee T, Orban J, Bryan P (1992) Thermodynamic analysis of the folding of the streptococcal protein G IgG-binding domains B1 and B2: why small proteins tend to have high denaturation temperatures. Biochemistry 31:3597–3603CrossRefGoogle Scholar
  2. Andersen NH, Neidigh JW, Harris SM, Lee GM, Liu Z, Tong H (1997) Extracting information from the temperature gradients of polypeptide NH chemical shifts. 1. The importance of conformational averaging. J Am Chem Soc 119:8547–8561CrossRefGoogle Scholar
  3. Baxter NJ, Williamson MP (1997) Temperature dependence of 1H chemical shifts in proteins. J Biomol NMR 9:359–369CrossRefGoogle Scholar
  4. Cierpicki T, Otlewski J (2001) Amide proton temperature coefficients as hydrogen bond indicators in proteins. J Biomol NMR 21:249–261CrossRefGoogle Scholar
  5. Cierpicki T, Zhukov I, Byrd RA, Otlewski J (2002) Hydrogen bonds in human ubiquitin reflected in temperature coefficients of amide protons. J Magn Reson 157:178–180ADSCrossRefGoogle Scholar
  6. Cordier F, Grzesiek S (2002) Temperature-dependence of protein hydrogen bond properties as studied by high-resolution NMR. J Mol Biol 715:739–752CrossRefGoogle Scholar
  7. Daley ME, Graether SP, Sykes BD (2004) Hydrogen bonding on the ice-binding face of a β-helical antifreeze protein indicated by amide proton NMR chemical shifts. Biochemistry 43:13012–13017CrossRefGoogle Scholar
  8. Findeisen M, Brand T, Berger S (2007) A 1H NMR thermometer suitable for cryoprobes. Magn Reson Chem 45:175–178CrossRefGoogle Scholar
  9. Gallagher T, Alexander P, Bryan P, Gilliland GL (1994) Two crystal structures of the B1 immunoglobulin-binding domain of Streptococcal protein G and comparison with NMR. Bichemistry 33:4721–4729CrossRefGoogle Scholar
  10. Joshi MF, Sidhu G, Nielsen JE, Brayer GD, Withers SG, McIntosh LP (2001) Dissecting the electrostatic interactions and pH-dependent activity of a family 11 glucosidase. Biochemistry 40:10115–10139CrossRefGoogle Scholar
  11. Ohnishi M, Urry DW (1969) Temperature dependence of amide proton chemical shifts: the secondary structures of gramicidin S and valinomycin. Biochem Biophys Res Comm 36:194–202CrossRefGoogle Scholar
  12. Schwarzinger S, Kroon GJA, Foss TR, Chung J, Wright PE, Dyson HJ (2001) Sequence-dependent correction of random coil NMR chemical shifts. J Am Chem Soc 123:2970–2978CrossRefGoogle Scholar
  13. Tomlinson JH, Ullah S, Hansen PE, Williamson MP (2009) Characterization of salt bridges to lysines in the protein G B1 domain. J Am Chem Soc 131:4674–4684CrossRefGoogle Scholar
  14. Tomlinson JH, Craven CJ, Williamson MP, Pandya MJ (2010a) Dimerization of protein G B1 domain at low pH: a conformational switch caused by loss of a single hydrogen bond. Proteins: Struct Funct Bioinf 78:1652–1661CrossRefGoogle Scholar
  15. Tomlinson JH, Green VL, Baker PJ, Williamson MP (2010b) Structural origins of pH-dependent chemical shifts in protein G. Proteins: Struct Funct Bioinf 78:3000–3016CrossRefGoogle Scholar
  16. Tunnicliffe RB, Waby JL, Williams RJ, Williamson MP (2005) An experimental investigation of conformational fluctuations in proteins G and L. Structure 13:1677–1684CrossRefGoogle Scholar
  17. Williamson MP, Craven CJ (2009) Automated protein structure calculation from NMR data. J Biomol NMR 43:131–143CrossRefGoogle Scholar
  18. Williamson MP, Waltho JP (1992) Peptide structure from NMR. Chem Soc Rev 21:227–236CrossRefGoogle Scholar
  19. Wishart DS, Bigam CG, Yao J, Abildgaard F, Dyson HJ, Oldfield E, Markley JL, Sykes BD (1995) 1H, 13C and 15N chemical shift referencing in biomolecular NMR. J Biomol NMR 6:135–140CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  1. 1.Department of Molecular Biology and BiotechnologyUniversity of SheffieldSheffieldUK
  2. 2.Institute of Molecular and Cellular BiologyUniversity of LeedsLeedsUK

Personalised recommendations