title: Angular dependences of perpendicular and parallel mode electron paramagnetic resonance of oxidized beef heart cytochrome c oxidase.
abstract: Cytochrome c oxidase catalyzes the reduction of oxygen to water with a concomitant conservation of energy in the form of a transmembrane proton gradient. The enzyme has a catalytic site consisting of a binuclear center of a copper ion and a heme group. The spectroscopic parameters of this center are unusual. The origin of broad electron paramagnetic resonance (EPR) signals in the oxidized state at rather low resonant field, the so-called g' = 12 signal, has been a matter of debate for over 30 years. We have studied the angular dependence of this resonance in both parallel and perpendicular mode X-band EPR in oriented multilayers containing cytochrome c oxidase to resolve the assignment. The "slow" form and compounds formed by the addition of formate and fluoride to the oxidized enzyme display these resonances, which result from transitions between states of an integer-spin multiplet arising from magnetic exchange coupling between the five unpaired electrons of high spin Fe(III) heme a(3) and the single unpaired electron of Cu(B). The first successful simulation of similar signals observed in both perpendicular and parallel mode X-band EPR spectra in frozen aqueous solution of the fluoride compound of the closely related enzyme, quinol oxidase or cytochrome bo(3), has been reported recently (Oganesyan et al., 1998, J. Am. Chem. Soc. 120:4232-4233). This suggested that the exchange interaction between the two metal ions of the binuclear center is very weak (|J| approximately 1 cm(-1)), with the axial zero-field splitting (D approximately 5 cm(-1)) of the high-spin heme dominating the form of the ground state. We show that this model accounts well for the angular dependences of the X-band EPR spectra in both perpendicular and parallel modes of oriented multilayers of cytochrome c oxidase derivatives and that the experimental results are inconsistent with earlier schemes that use exchange coupling parameters of several hundred wavenumbers.
authors.affiliation: School of Biological and Medical Sciences, University of St. Andrews, St. Andrews, Fife KY16 9AL, Scotland.
authors.forename: D J
authors.initials: DJ
authors.lastname: Hunter
authors.forename: V S
authors.initials: VS
authors.lastname: Oganesyan
authors.forename: J C
authors.initials: JC
authors.lastname: Salerno
authors.forename: C S
authors.initials: CS
authors.lastname: Butler
authors.forename: W J
authors.initials: WJ
authors.lastname: Ingledew
authors.forename: A J
authors.initials: AJ
authors.lastname: Thomson
mesh_terms: D000818:Animals; D002417:Cattle; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D003576:Electron Transport Complex IV / Q000737:chemistry*; D006418:Heme / Q000032:analysis; D007425:Intracellular Membranes / Q000201:enzymology; D008929:Mitochondria, Heart / Q000201:enzymology*; D010084:Oxidation-Reduction; D013367:Submitochondrial Particles / Q000201:enzymology; D013816:Thermodynamics
chemical_list: D006418:Heme; D003576:Electron Transport Complex IV