title: Electron transfer in ruthenium-modified spinach plastocyanin mutants.
abstract: Four site-directed mutants of spinach plastocyanin, Pc(Leu12His), Pc(Leu15His), Pc(Thr79His), and Pc(Lys81His), have been modified by covalent attachment of a photoactive [Ru(bpy)2(im)]2+ complex at the surface-exposed histidine residues. The Pc-Ru complexes were characterized with optical absorption, CD, and EPR spectroscopy and their spectra were found to be similar to the unmodified proteins except in the case of the Pc(Leu12His) mutant which lost the Cu ion irreversibly during the Ru modification. Electron transfer (ET) within the other Pc-Ru complexes was studied with time-resolved optical spectroscopy, using an external-quencher approach. The fully reduced [Cu(I), Ru(II)] proteins were photoexcited and subsequently oxidized by an external quencher, [Ru(NH3)6]Cl3, forming the [Cu(I), Ru(III)] proteins. This was followed by an internal ET from Cu(I) to Ru(III). The rates of the internal ET reactions exhibit an exponential dependence on metal-to-metal separation, with a decay factor of 1.1 A-1. From a temperature-dependence study of the Ru-modified Pc(Lys81His) protein, a reorganization energy for the Cu-to-Ru ET reaction of 1.2 eV was determined. In this analysis it was found necessary to include an appreciable temperature dependence in the driving force of the ET reaction.
authors.affiliation: Lundberg Laboratory, Göteborg University and Chalmers University of Technology, Medicinaregatan 9C, Göteborg, S-413 90, Sweden. karl.sigfridsson@nordicsynthesis.se
authors.forename: K
authors.initials: K
authors.lastname: Sigfridsson
authors.forename: M
authors.initials: M
authors.lastname: Ejdebäck
authors.forename: M
authors.initials: M
authors.lastname: Sundahl
authors.lastname: Hansson
mesh_terms: D003300:Copper / Q000737:chemistry / Q000378:metabolism; D004579:Electron Transport / Q000502:physiology*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D010782:Photolysis; D010970:Plastocyanin / Q000737:chemistry* / Q000235:genetics; D017434:Protein Structure, Tertiary; D017975:Ruthenium Compounds / Q000378:metabolism / Q000494:pharmacology*; D013057:Spectrum Analysis; D018724:Spinacia oleracea / Q000502:physiology*; D013696:Temperature
chemical_list: D017975:Ruthenium Compounds; D003300:Copper; D010970:Plastocyanin