Determination of the rate constant of self-exchange of the oxygen O2/O2- couple in water by 18O/16O isotope marking

Lind, Johan; Shen, X.; Merenyi, Gabar; Jonsson, B.Ö.

doi:10.1021/ja00201a078

Cited by 68 publications

(64 citation statements)

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“…In aqueous media the total reorganization energy, λ, has been determined to be 1.97 eV. 51 In a calculation by Fukuzumi, based on thermodynamic data available, a Cu I −O 2 species where dioxygen is hypothetically bound to copper(I) prior to electrontransfer), the internal (inner-sphere) electron-transfer from the copper(I) to the O 2 to give a cupric-superoxide product (that species which is observed) gives a calculated λ (total) value of 1.74 eV. 47b A related calculation indicates that free dioxygen binds to a porphryinate-cobalt(II) complex to give the Co(III)- superoxide species with λ = 1.89 eV.…”

Section: Resultsmentioning

confidence: 99%

Peroxo and Superoxo Moieties Bound to Copper Ion: Electron-Transfer Equilibrium with a Small Reorganization Energy

et al. 2016

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Oxygenation of [Cu2(UN-O−)(DMF)]2+ (1), a structurally characterized dicopper Robin–Day class I mixed-valent Cu(II)Cu(I) complex, with UN-O− as a binucleating ligand and where dimethylformamide (DMF) binds to the Cu(II) ion, leads to a superoxo-dicopper(II) species [CuII2(UN-O−)(O2•−)]2+ (2). The formation kinetics provide that kon = 9 × 10−2 M−1 s−1 (−80 °C), ΔH‡ = 31.1 kJ mol−1 and ΔS‡ = −99.4 J K−1 mol−1 (from −60 to −90 °C data). Complex 2 can be reversibly reduced to the peroxide species [CuII2(UN-O−)(O22−)]+ (3), using varying outer-sphere ferrocene or ferrocenium redox reagents. A Nernstian analysis could be performed by utilizing a monodiphenylamine substituted ferrocenium salt to oxidize 3, leading to an equilibrium mixture with Ket = 5.3 (−80 °C); a standard reduction potential for the superoxo–peroxo pair is calculated to be E° = +130 mV vs SCE. A literature survey shows that this value falls into the range of biologically relevant redox reagents, e.g., cytochrome c and an organic solvent solubilized ascorbate anion. Using mixed-isotope resonance Raman (rRaman) spectroscopic characterization, accompanied by DFT calculations, it is shown that the superoxo complex consists of a mixture of μ-1,2- (21,2) and μ-1,1- (21,1) isomers, which are in rapid equilibrium. The electron transfer process involves only the μ-1,2-superoxo complex [CuII2(UN-O−)(μ-1,2-O2•−)]2+ (21,2) and μ-1,2-peroxo structures [CuII2(UN-O−)(O22−)]+ (3) having a small bond reorganization energy of 0.4 eV (λin). A stopped-flow kinetic study results reveal an outer-sphere electron transfer process with a total reorganization energy (λ) of 1.1 eV between 21,2 and 3 calculated in the context of Marcus theory.

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Section: Resultsmentioning

confidence: 99%

Peroxo and Superoxo Moieties Bound to Copper Ion: Electron-Transfer Equilibrium with a Small Reorganization Energy

et al. 2016

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“…The positively charged ε-amino group of Lys265 might activate oxygen by increasing the 1-electron reduction potential for the O 2 /O 2 •- couple (E° = −160 mV vs NHE), similar to the effect observed with cations in model studies (35). However, the kinetics observed for the self-exchange reaction between oxygen and superoxide anion indicate that λ out constitutes the major energy barrier in the 1-electron reduction of oxygen (36). Importantly, the region above the si -face of FAD in MSOX contains a pair of water molecules that are sandwiched between flavin N(5) and the positively charged ε-amino group of Lys265 (Figure 1).…”

Section: Discussionmentioning

confidence: 99%

Identification of the Oxygen Activation Site in Monomeric Sarcosine Oxidase: Role of Lys265 in Catalysis

2008

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Monomeric sarcosine oxidase (MSOX) catalyzes the oxidation of N-methylglycine and contains covalently bound FAD that is hydrogen bonded at position N(5) to Lys265 via a bridging water. Lys265 is absent in the homologous but oxygen-unreactive FAD site in heterotetrameric sarcosine oxidase. Isolated preparations of Lys265 mutants contain little or no flavin but can be covalently reconstituted with FAD. Mutation of Lys265 to a neutral residue (Ala, Gln, Met) causes a 6000-to 9000-fold decrease in apparent turnover rate whereas a 170-fold decrease is found with Lys265Arg. Substitution of Lys265 with Met or Arg causes only a modest decrease in the rate of sarcosine oxidation (9.0-or 3.8-fold, respectively), as judged by reductive half-reaction studies which show that the reactions proceed via an initial enzyme•sarcosine charge transfer complex and a novel spectral intermediate not detected with wild-type MSOX. Oxidation of reduced wild-type MSOX (k = 2.83 × 10 5 M −1 s −1 ) is more than 1000-fold faster than observed for the reaction of oxygen with free reduced flavin. Mutation of Lys265 to a neutral residue causes a dramatic 8000-fold decrease in oxygen reactivity whereas a 250-fold decrease is observed with Lys265Arg. The results provide definitive evidence for Lys265 as the site of oxygen activation and show that a single positively charged amino acid residue is entirely responsible for the rate acceleration observed with wild-type enzyme. Significantly, the active sites for sarcosine oxidation and oxygen reduction are located on opposite faces of the flavin ring.The reduction of oxygen to hydrogen peroxide by free reduced flavin is thermodynamically favorable but kinetically slow because the 2-electron reduction of triplet oxygen by a diamagnetic organic molecule is spin-forbidden. In fact, the 2-electron reduction of oxygen by reduced flavin proceeds via an initial 1-electron transfer step that generates a flavin radical/ superoxide anion radical pair in a spin-allowed but energetically unfavorable, rate-determining reaction. The term oxygen activation is used in reference to the accelerated rates of oxygen reduction observed with flavoprotein oxidases and other enzymes that reduce molecular oxygen (1,2). A series of elegant studies by Klinman and Roth identified His516 as the site of oxygen activation in the flavoenzyme glucose oxidase and showed that the reaction required the protonated form of this residue (2-4). Surprisingly little is, however, currently known about the detailed mechanism of oxygen activation by other flavoprotein oxidases, especially regarding the specific role of active site residues in these reactions.

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“…4 (39) but may be as low as 11 kcal⅐mol Ϫ1 according to recent quantum treatments (40). The former value is used here to estimate out ϭ 30 kcal⅐mol Ϫ1 (1.3 eV) from the difference Ϫ in (39). This finding indicates that reorganization of the surrounding medium presents a major energy cost for electron transfer to O 2 .…”

Section: [3]mentioning

confidence: 92%

Catalysis of electron transfer during activation of O ₂ by the flavoprotein glucose oxidase

Roth

Klinman

2002

Proc. Natl. Acad. Sci. U.S.A.

170

281

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Two prototropic forms of glucose oxidase undergo aerobic oxidation reactions that convert FADH ؊ to FAD and form H2O2 as a product. Limiting rate constants of kcat͞KM(O2) ‫؍‬ (5.7 ؎ 1.8) ؋ 10 2 M ؊1 ⅐s ؊1 and kcat͞KM(O2) ‫؍‬ (1.5 ؎ 0.3) ؋ 10 6 M ؊1 ⅐s ؊1 are observed at high and low pH, respectively. Reactions exhibit oxygen-18 kinetic isotope effects but no solvent kinetic isotope effects, consistent with mechanisms of rate-limiting electron transfer from flavin to O2. Site-directed mutagenesis studies reveal that the pH dependence of the rates is caused by protonation of a highly conserved histidine in the active site. Temperature studies (283-323 K) indicate that protonation of His-516 results in a reduction of the activation energy barrier by 6.0 kcal⅐mol ؊1 (0.26 eV). Within the context of Marcus theory, catalysis of electron transfer is attributed to a 19-kcal⅐mol ؊1 (0.82 eV) decrease in the reorganization energy and a much smaller 2.2-kcal⅐mol ؊1 (0.095 eV) enhancement of the reaction driving force. An explanation is advanced that is based on changes in outer-sphere reorganization as a function of pH. The active site is optimized at low pH, but not at high pH or in the H516A mutant where rates resemble the uncatalyzed reaction in solution. F lavins are highly versatile enzyme cofactors that undergo electron and proton-coupled electron transfer reactions (1-3). As a result, flavoenzymes are involved in an array of chemical and photochemical processes from COH oxidations (4) to electron transport (5) to repair of cross-linked DNA (6). Glucose oxidase (GO) is a homodimeric protein found predominantly in fungi (National Center for Biotechnology Information, www.ncbi.nlm.nih.gov). Each protein subunit contains an equivalent of noncovalently bound FAD Ϸ15 Å below the surface (7). GO mediates net hydride transfer from the anomeric COH bond of glucose to FAD in the reductive half-reaction (8, 9) and the oxidation of reduced cofactor (FADH Ϫ ) by O 2 in the oxidative half-reaction, forming H 2 O 2 as a product. All evidence points toward a rate-limiting electron transfer step during FADH Ϫ oxidation as shown in Eq. 1 (10). This reaction involves the transfer of negative charge from cofactor to superoxide ion with no net change in charge at the active site.To date, most mechanistic studies of O 2 activation have focused on metalloenzymes. In such reactions, electron transfer and electrostatic stabilization often occur within a single step (ref. 11 and references therein), causing rates to approach the diffusion limit (12). Additionally, ligands control metal coordination geometries and modulate redox potentials, thereby tuning reactivity toward O 2 (13). Enzymes that use organic cofactors do not enjoy such advantages, but may use specialized protein environments to help overcome the kinetic and thermodynamic barriers associated with activation of O 2 .The physical characterization of the protein dielectric is an area of growing interest (14-18) and may provide the key to understanding how proteins like GO facilita...

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Determination of the rate constant of self-exchange of the oxygen O2/O2- couple in water by 18O/16O isotope marking

Cited by 68 publications

References 1 publication

Peroxo and Superoxo Moieties Bound to Copper Ion: Electron-Transfer Equilibrium with a Small Reorganization Energy

Peroxo and Superoxo Moieties Bound to Copper Ion: Electron-Transfer Equilibrium with a Small Reorganization Energy

Identification of the Oxygen Activation Site in Monomeric Sarcosine Oxidase: Role of Lys265 in Catalysis

Catalysis of electron transfer during activation of O ₂ by the flavoprotein glucose oxidase

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Determination of the rate constant of self-exchange of the oxygen O2/O2- couple in water by 18O/16O isotope marking

Cited by 68 publications

References 1 publication

Peroxo and Superoxo Moieties Bound to Copper Ion: Electron-Transfer Equilibrium with a Small Reorganization Energy

Peroxo and Superoxo Moieties Bound to Copper Ion: Electron-Transfer Equilibrium with a Small Reorganization Energy

Identification of the Oxygen Activation Site in Monomeric Sarcosine Oxidase: Role of Lys265 in Catalysis

Catalysis of electron transfer during activation of O 2 by the flavoprotein glucose oxidase

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Catalysis of electron transfer during activation of O ₂ by the flavoprotein glucose oxidase