Magnetic state of the a3 center of cytochrome c oxidase and some of its derivatives

Abstract

The temperature dependence of the magnetic susceptibility was used to investigate the nature of the coupling between cytochrome alpha 3 and CuB in resting and oxidized cyanide- and formate-bound cytochrome oxidase. Resting and formate-bound enzymes were found to have strong antiferromagnetic coupling with an S = 5/2 cytochrome alpha 3, results that were independent of the dispersing detergent and the enzyme isolation method. The cyanide-bound enzyme was heterogeneous, with a minor fraction showing intermediate strength antiferromagnetic coupling. The magnitude of this coupling was independent of the enzyme isolation method and depended moderately on the identity of the dispersing detergent. The major fraction of the cyanide-bound enzyme had a lowest energy state of Ms = 0. The coupling constant for this fraction did not depend on the isolation technique or on the identity of the dispersing detergent. The use of glucose-glucose oxidase to deoxygenate samples influenced the susceptibility behavior of some preparations of both the resting and formate-bound enzymes, with results indicating an S = 3/2 cytochrome alpha 3 in the resting enzyme samples. Retention of a 417-nm Soret band for formate-bound enzyme concomitant with peroxide-induced changes in susceptibility behavior indicates different sites of enzyme interactions for the formate ion and hydrogen peroxide.

Magnetization of fast and slow oxidized cytochrome c oxidase

“…In the current study, the increased information of the multifield saturation magnetization method results in a clear distinction between the fast and slow forms of oxidized cytochrome c oxidase. A previous susceptibility study was unable to distinguish these forms of the enzyme (Barnes et al, 1991).…”

Angular Dependences of Perpendicular and Parallel Mode Electron Paramagnetic Resonance of Oxidized Beef Heart Cytochrome c Oxidase

“…Theoretical models based on both the strong and weak coupling limits have also been used to interpret Mössbauer and magnetic susceptibility data on cytochrome c oxidase (Moss et al, 1978;Dunham et al, 1983;Kent et al, 1983;Tweedle et al, 1978;Day et al, 1993;Barnes et al, 1991;Rusnak et al, 1987) as well as on heme-copper-bridged assemblies pertinent to cytochrome c oxidase (Kauffmann et al, 1997). Mössbauer spectra of the a 3 -Cu B site of oxidized Thermus thermophilus c 1 aa 3 could be simulated equally well in terms of a weak (J Ϸ 1 cm Ϫ1 ) or a strong (J Ͼ 7 cm Ϫ1 ) coupling (Rusnak et al, 1987).…”

Intermolecular Interactions and Intramolecular Couplings of Binuclear Porphyrin Models for Cytochrome c Oxidase

“…12,13 However, in the slow form, the magnetic coupling and zero-field-splitting parameters are quantitatively different from those of the fast enzyme and result in the observation of an active EPR spectrum. 7,14,15 Dual-mode X-band EPR spectra of fluoride cytochrome bo 3 show broad, fast relaxing features with a similar pattern of bands: a weak derivative signal below 100 mT (g ≈ 12) accompanied by a broad band in the 200−250 mT region (g ≈ 3.2). 10,16−18 Simulations of dual-mode spectra indicated that the metal ions were weakly coupled by an anisotropic exchange interaction of |J| ≈ 1 cm −1 .…”

“…In the oxidized C c O, fast and slow forms are defined according to the rate of reaction of the enzyme with exogenous anionic ligands such as fluoride (F – ), cyanide (CN – ), and formate (HCOO – ). , Both forms of the enzyme have been proposed to consist of the Fe III –X–Cu II moiety, in which X represents a bridging ligand that modulates the coupling between two metal ions. , Naturally occurring X groups, which have been proposed, include oxide (O 2– ), hydroxide (OH – ), cyanide (CN – ), formate (HCOO – ), chloride (Cl – ), fluoride (F – ), and an imidazolate group . Mössbauer spectroscopic measurements of Paracoccus denitrificans and bovine heart C c O revealed that high-spin Fe 3+ ( S = 5/2) was strongly coupled with Cu 2+ ( S = 1/2), resulting in no EPR signal in the fast form of the enzyme. , However, in the slow form, the magnetic coupling and zero-field-splitting parameters are quantitatively different from those of the fast enzyme and result in the observation of an active EPR spectrum. ,, Dual-mode X-band EPR spectra of fluoride cytochrome bo 3 show broad, fast relaxing features with a similar pattern of bands: a weak derivative signal below 100 mT ( g ≈ 12) accompanied by a broad band in the 200–250 mT region ( g ≈ 3.2). ,− Simulations of dual-mode spectra indicated that the metal ions were weakly coupled by an anisotropic exchange interaction of | J | ≈ 1 cm –1 .…”

“…4,5 Both forms of the enzyme have been proposed to consist of the Fe III −X−Cu II moiety, in which X represents a bridging ligand that modulates the coupling between two metal ions. 6,7 Naturally occurring X groups, which have been proposed, include oxide (O 2− ), 8 hydroxide (OH − ), 8 cyanide (CN − ), 9 formate (HCOO − ), 10 chloride (Cl − ), 10 fluoride (F − ), 10 and an imidazolate group. 11 Mossbauer spectroscopic measurements of Paracoccus denitrif icans and bovine heart CcO revealed that high-spin Fe 3+ (S = 5/2) was strongly coupled with Cu 2+ (S = 1/2), resulting in no EPR signal in the fast form of the enzyme.…”