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Direct access to the cooperative substructure of proteins and the protein ensemble via cold denaturation

Nature Structural & Molecular Biology volume 11, pages 352–357 (2004)Cite this article

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Abstract

The modern view of protein thermodynamics predicts that proteins undergo cold-induced unfolding. Unfortunately, the properties of proteins and water conspire to prevent the detailed observation of this fundamental process. Here we use protein encapsulation to allow cold denaturation of the protein ubiquitin to be monitored by high-resolution NMR at temperatures approaching −35 °C. The cold-induced unfolding of ubiquitin is found to be highly noncooperative, in distinct contrast to the thermal melting of this and other proteins. These results demonstrate the potential of cold denaturation as a means to dissect the cooperative substructures of proteins and to provide a rigorous framework for testing statistical thermodynamic treatments of protein stability, dynamics and function.

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Figure 1: Implications of the temperature dependence of protein stability.
Figure 2: Schematic illustration drawn to scale of the dimensions of the AOT-encapsulated ubiquitin reverse micelle system at a W0 of 10.
Figure 3: Cold-induced unfolding of encapsulated ubiquitin by high-resolution solution NMR spectroscopy.
Figure 4: Slow interconversion between native and cold-induced states of ubiquitin.
Figure 5: The progressive cold-induced structural transitions of ubiquitin.
Figure 6: Statistical thermodynamic analysis of the cold denaturation of ubiquitin using the COREX algorithm18.

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  • 07 March 2004

    Added note and updated figure online, updated PDF

Notes

  1. *Note: In the version of this article initially published online, the temperature for the far right-hand panels of Figure 3a,b was incorrectly labeled during the production process. The correct temperature should be −30 °C. We apologize for any inconvenience this may have caused. This mistake has been corrected for the HTML and print versions of the article.

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Acknowledgements

This work was supported by grants from the US National Institutes of Health. We are grateful to K.A. Sharp and S.W. Englander for helpful discussion and to S. Whitten and P.F. Flynn for technical assistance.

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Authors and Affiliations

  1. Johnson Research Foundation and Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, 19104-6059, Pennsylvania, USA

    Charles R Babu & A Joshua Wand

  2. Department of Human Biological Chemistry and Genetics and the Sealy Center for Structural Biology, University of Texas Medical Branch, Galveston, 77555-1055, Texas, USA

    Vincent J Hilser

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Correspondence to A Joshua Wand.

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Babu, C., Hilser, V. & Wand, A. Direct access to the cooperative substructure of proteins and the protein ensemble via cold denaturation. Nat Struct Mol Biol 11, 352–357 (2004). https://doi.org/10.1038/nsmb739

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