title: Mutations in the docking site for cytochrome c on the Paracoccus heme aa3 oxidase. Electron entry and kinetic phases of the reaction.
abstract: Introducing site-directed mutations in surface-exposed residues of subunit II of the heme aa3 cytochrome c oxidase of Paracoccus denitrificans, we analyze the kinetic parameters of electron transfer from reduced horse heart cytochrome c. Specifically we address the following issues: (a) which residues on oxidase contribute to the docking site for cytochrome c, (b) is an aromatic side chain required for electron entry from cytochrome c, and (c) what is the molecular basis for the previously observed biphasic reaction kinetics. From our data we conclude that tryptophan 121 on subunit II is the sole entry point for electrons on their way to the CuA center and that its precise spatial arrangement, but not its aromatic nature, is a prerequisite for efficient electron transfer. With different reaction partners and experimental conditions, biphasicity can always be induced and is critically dependent on the ionic strength during the reaction. For an alternative explanation to account for this phenomenon, we find no evidence for a second cytochrome c binding site on oxidase.
authors.affiliation: Molecular Genetics, Institute of Biochemistry, Johann-Wolfgang-Goethe Universität, Frankfurt, Germany.
authors.forename: Viktoria
authors.initials: V
authors.lastname: Drosou
authors.forename: Francesco
authors.initials: F
authors.lastname: Malatesta
authors.forename: Bernd
authors.initials: B
authors.lastname: Ludwig
mesh_terms: D001665:Binding Sites; D003574:Cytochrome c Group / Q000235:genetics / Q000378:metabolism*; D004579:Electron Transport / Q000502:physiology; D003576:Electron Transport Complex IV / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010231:Paracoccus denitrificans / Q000201:enzymology*; D011485:Protein Binding; D021122:Protein Subunits; D014364:Tryptophan / Q000378:metabolism*
chemical_list: D003574:Cytochrome c Group; D021122:Protein Subunits; D014364:Tryptophan; D003576:Electron Transport Complex IV