A copper protein and a cytochrome bind at the same site on bacterial cytochrome c peroxidase

Pauleta, S.R., Cooper, A., Nutley, M., Errington, N., Harding, S., Guerlesquin, F., Goodhew, C.F., Moura, I., Moura, J.J.G. and Pettigrew, G.W. (2004) A copper protein and a cytochrome bind at the same site on bacterial cytochrome c peroxidase. Biochemistry, 43(46), pp. 14566-14576. (doi:10.1021/bi0485833)

Full text not currently available from Enlighten.

Abstract

Pseudoazurin binds at a single site on cytochrome c peroxidase from Paracoccus pantotrophus with a Kd of 16.4 μM at 25 °C, pH 6.0, in an endothermic reaction that is driven by a large entropy change. Sedimentation velocity experiments confirmed the presence of a single site, although results at higher pseudoazurin concentrations are complicated by the dimerization of the protein. Microcalorimetry, ultracentrifugation, and 1H NMR spectroscopy studies in which cytochrome c550, pseudoazurin, and cytochrome c peroxidase were all present could be modeled using a competitive binding algorithm. Molecular docking simulation of the binding of pseudoazurin to the peroxidase in combination with the chemical shift perturbation pattern for pseudoazurin in the presence of the peroxidase revealed a group of solutions that were situated close to the electron-transferring heme with Cu−Fe distances of about 14 Å. This is consistent with the results of 1H NMR spectroscopy, which showed that pseudoazurin binds closely enough to the electron-transferring heme of the peroxidase to perturb its set of heme methyl resonances. We conclude that cytochrome c550 and pseudoazurin bind at the same site on the cytochrome c peroxidase and that the pair of electrons required to restore the enzyme to its active state after turnover are delivered one-by-one to the electron-transferring heme.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Nutley, Mrs Margaret and Cooper, Professor Alan
Authors: Pauleta, S.R., Cooper, A., Nutley, M., Errington, N., Harding, S., Guerlesquin, F., Goodhew, C.F., Moura, I., Moura, J.J.G., and Pettigrew, G.W.
College/School:College of Science and Engineering > School of Chemistry
Journal Name:Biochemistry
Publisher:American Chemical Society
ISSN:0006-2960
ISSN (Online):1520-4995
Published Online:29 October 2004

University Staff: Request a correction | Enlighten Editors: Update this record