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Splitting of the O-O bond at the heme-copper catalytic site of respiratory oxidases.

ABSTRACT: Heme-copper oxidases catalyze the four-electron reduction of O2 to H2O at a catalytic site that is composed of a heme group, a copper ion (CuB), and a tyrosine residue. Results from earlier experimental studies have shown that the O-O bond is cleaved simultaneously with electron transfer from a low-spin heme (heme a/b), forming a ferryl state (PR ; Fe4+=O2-, CuB2+-OH-). We show that with the Thermus thermophilus ba3 oxidase, at low temperature (10°C, pH 7), electron transfer from the low-spin heme b to the catalytic site is faster by a factor of ~10 (? ? 11 ?s) than the formation of the PR ferryl (? ?110 ?s), which indicates that O2 is reduced before the splitting of the O-O bond. Application of density functional theory indicates that the electron acceptor at the catalytic site is a high-energy peroxy state [Fe3+-O--O-(H+)], which is formed before the PR ferryl. The rates of heme b oxidation and PR ferryl formation were more similar at pH 10, indicating that the formation of the high-energy peroxy state involves proton transfer within the catalytic site, consistent with theory. The combined experimental and theoretical data suggest a general mechanism for O2 reduction by heme-copper oxidases.

SUBMITTER: Poiana F

PROVIDER: S-EPMC5473675 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Splitting of the O-O bond at the heme-copper catalytic site of respiratory oxidases.

Poiana Federica F von Ballmoos Christoph C Gonska Nathalie N Blomberg Margareta R A MRA Ädelroth Pia P Brzezinski Peter P

Science advances 20170616 6

Heme-copper oxidases catalyze the four-electron reduction of O2 to H2O at a catalytic site that is composed of a heme group, a copper ion (CuB), and a tyrosine residue. Results from earlier experimental studies have shown that the O-O bond is cleaved simultaneously with electron transfer from a low-spin heme (heme a/b), forming a ferryl state (PR ; Fe4+=O2-, CuB2+-OH-). We show that with the ...[more]

PMID: 28630929

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Project description:The route of O₂to and from the high-spin heme in heme-copper oxidases has generally been believed to emulate that of carbon monoxide (CO). Time-resolved and stationary infrared experiments in our laboratories of the fully reduced CO-bound enzymes, as well as transient optical absorption saturation kinetics studies as a function of CO pressure, have provided strong support for CO binding to CuB⁺ on the pathway to and from the high-spin heme. The presence of CO on CuB⁺ suggests that O₂binding may be compromised in CO flow-flash experiments. Time-resolved optical absorption studies show that the rate of O₂and NO binding in the bovine enzyme (1 × 10⁸M⁻¹s⁻¹) is unaffected by the presence of CO, which is consistent with the rapid dissociation (t½ = 1.5μs) of CO from CuB⁺. In contrast, in Thermus thermophilus (Tt) cytochrome ba3 the O₂and NO binding to heme a3 slows by an order of magnitude in the presence of CO (from 1 × 10⁹ to 1 × 10⁸M⁻¹s⁻¹), but is still considerably faster (~10μs at 1atm O₂) than the CO off-rate from CuB in the absence of O₂(milliseconds). These results show that traditional CO flow-flash experiments do not give accurate results for the physiological binding of O₂and NO in Tt ba3, namely, in the absence of CO. They also raise the question whether in CO flow-flash experiments on Tt ba3 the presence of CO on CuB⁺ impedes the binding of O₂to CuB⁺ or, if O₂does not bind to CuB⁺ prior to heme a3, whether the CuB⁺-CO complex sterically restricts access of O₂to the heme. Both possibilities are discussed, and we argue that O₂binds directly to heme a3 in Tt ba3, causing CO to dissociate from CuB⁺ in a concerted manner through steric and/or electronic effects. This would allow CuB⁺ to function as an electron donor during the fast (5μs) breaking of the OO bond. These results suggest that the binding of CO to CuB⁺ on the path to and from heme a3 may not be applicable to O₂and NO in all heme-copper oxidases. This article is part of a Special Issue entitled: Vibrational spectroscopies and bioenergetic systems.

Dataset Information

Splitting of the O-O bond at the heme-copper catalytic site of respiratory oxidases.

Publications

Splitting of the O-O bond at the heme-copper catalytic site of respiratory oxidases.

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