A Caltech Library Service

Effect of Redox State on the Folding Free Energy of a Thermostable Electron-Transfer Metalloprotein: The Cu_A Domain of Cytochrome Oxidase from Thermus thermophilus

Wittung-Stafshede, Pernilla and Malmström, Bo G. and Sanders, Donita and Fee, James A. and Winkler, Jay R. and Gray, Harry B. (1998) Effect of Redox State on the Folding Free Energy of a Thermostable Electron-Transfer Metalloprotein: The Cu_A Domain of Cytochrome Oxidase from Thermus thermophilus. Biochemistry, 37 (9). pp. 3172-3177. ISSN 0006-2960. doi:10.1021/bi972901z.

Full text is not posted in this repository. Consult Related URLs below.

Use this Persistent URL to link to this item:


The unfolding of the Cu_A domain of cytochrome oxidase from the thermophilic bacterium Thermus thermophilus, induced by guanidine hydrochloride (GuHCl)^1 at different temperatures, has been monitored by CD as well by electronic absorption (with the oxidized protein) and by fluorescence (with the reduced protein). The same unfolding curves were obtained with the different methods, providing evidence for a two-state model for the unfolding equilibrium. This was also supported by the shape of the unfolding equilibrium curves and by the observed refolding of the unfolded, oxidized protein on dilution of the denaturant. The oxidized protein cannot be unfolded by GuHCl at room temperature, and it was found to be thermally very stable as well, since, even in the presence of 7 M GuHCl, it is not fully unfolded until above 80 °C. For the reduced protein at room temperature, the unfolding equilibrium curve yielded a folding free energy of −65 kJ/mol. The corresponding value for the oxidized protein (−85 kJ/mol) could be estimated indirectly from a thermodynamic cycle connecting the folded and unfolded forms in both oxidation states and the known reduction potentials of the metal site in the folded and unfolded states; the potential is increased on unfolding, consistent with the higher folding stability of the oxidized form. The difference in folding stability between the oxidized and reduced proteins (20 kJ/mol) is exceptionally high, and this is ascribed to the unique structure of the dinuclear CuA site. The unfolded, reduced protein was found to refold partially on oxidation with ferricyanide.

Item Type:Article
Related URLs:
URLURL TypeDescription
Wittung-Stafshede, Pernilla0000-0003-1058-1964
Winkler, Jay R.0000-0002-4453-9716
Gray, Harry B.0000-0002-7937-7876
Additional Information:© 1998 American Chemical Society. Received November 26, 1997. Publication Date (Web): February 14, 1998. This work was supported by the National Institutes of Health (Grant DK19038), the National Science Foundation (Grant MCB9630465), and the Swedish Natural Science Research Council. P.W.-S. is the recipient of a postdoctoral fellowship from Swedish Technical Research Council. We thank Dr. Stephen L. Mayo for the use of the Aviv spectropolarimeter.
Funding AgencyGrant Number
Swedish Natural Science Research CouncilUNSPECIFIED
Swedish Technical Research CouncilUNSPECIFIED
Issue or Number:9
Record Number:CaltechAUTHORS:20171120-144202285
Persistent URL:
Official Citation:Effect of Redox State on the Folding Free Energy of a Thermostable Electron-Transfer Metalloprotein:  The CuA Domain of Cytochrome Oxidase from Thermus thermophiles. Pernilla Wittung-Stafshede, Bo G. Malmström, Donita Sanders, James A. Fee, Jay R. Winkler, and Harry B. Gray. Biochemistry 1998 37 (9), 3172-3177. DOI: 10.1021/bi972901z
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:83359
Deposited By: Tony Diaz
Deposited On:20 Nov 2017 22:51
Last Modified:15 Nov 2021 19:57

Repository Staff Only: item control page