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European Biophysics Journal

, Volume 35, Issue 4, pp 363–366 | Cite as

Combined NMR-observation of cold denaturation in supercooled water and heat denaturation enables accurate measurement of ΔC p of protein unfolding

  • Thomas Szyperski
  • Jeffrey L. Mills
  • Dieter Perl
  • Jochen Balbach
Biophysics Letter

Abstract

Cold and heat denaturation of the double mutant Arg 3→Glu/Leu 66→Glu of cold shock protein Csp of Bacillus caldolyticus was monitored using 1D 1H NMR spectroscopy in the temperature range from −12°C in supercooled water up to +70°C. The fraction of unfolded protein, f u, was determined as a function of the temperature. The data characterizing the unfolding transitions could be consistently interpreted in the framework of two-state models: cold and heat denaturation temperatures were determined to be −11°C and 39°C, respectively. A joint fit to both cold and heat transition data enabled the accurate spectroscopic determination of the heat capacity difference between native and denatured state, ΔC p of unfolding. The approach described in this letter, or a variant thereof, is generally applicable and promises to be of value for routine studies of protein folding.

Keywords

Protein cold denaturation Cold shock protein Protein folding thermodynamics Supercooled water 

Notes

Acknowledgements

This work was supported by a Research Innovation Award of the Research Corporation (to T.S.) and the National Science Foundation (MCB 0416899 to T.S.), grants of the Deutsche Forschungsgemeinschaft (DFG 1821/2-1 and DFG 1821/3-1 to J. B.), and INTAS 2001-2347 (to J.B.). We thank M. Zeeb and F. X. Schmid for helpful discussions.

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Copyright information

© EBSA 2005

Authors and Affiliations

  • Thomas Szyperski
    • 1
  • Jeffrey L. Mills
    • 1
    • 2
  • Dieter Perl
    • 3
    • 4
  • Jochen Balbach
    • 4
    • 5
  1. 1.Department of ChemistryThe State University of New YorkBuffaloUSA
  2. 2.E633B School of MedicineCase Western Reserve UniversityClevelandUSA
  3. 3.Biotechnology DevelopmentNovartis Pharma AGBaselSwitzerland
  4. 4.Laboratorium für BiochemieUniversität BayreuthBayreuthGermany
  5. 5.Fachbereich Physik, Fachgruppe BiophysikMartin-Luther-Universität Halle-WittenbergHalle (Saale)Germany

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