Oxygen activation by a mixed-valent, diiron(II/III) cluster in the glycol cleavage reaction catalyzed by myo-inositol oxygenase

Gang Xing, Eric W. Barr, Yinghui Diao, Lee M. Hoffart, Kumble Sandeep Prabhu, Ryan J. Arner, C. Channa Reddy, Carsten Krebs, Joseph M. Bollinger, Jr.

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

myo-Inositol oxygenase (MIOX) catalyzes the ring-cleaving, four-electron oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate (myo-inositol, MI) to D-glucuronate (DG). The preceding paper [Xing, G., Hoffart, L. M., Diao, Y., Prabhu, K. S., Arner, R. J., Reddy, C. C., Krebs, C., and Bollinger, J. M., Jr. (2006) Biochemistry 45, 5393-5401] demonstrates by Mössbauer and electron paramagnetic resonance (EPR) spectroscopies that MIOX can contain a non-heme dinuclear iron cluster, which, in its mixed-valent (II/III) and fully oxidized (III/III) states, is perturbed by binding of MI in a manner consistent with direct coordination. In the study presented here, the redox form of the enzyme that activates O2 has been identified. L-Cysteine, which was previously reported to accelerate turnover, reduces the fully oxidized enzyme to the mixed-valent form, and O2, the cosubstrate, oxidizes the fully reduced form to the mixed-valent form with a stoichiometry of one per O 2, Both observations implicate the mixed-valent, diiron-(II/III) form of the enzyme as the active state. Stopped-flow absorption and freeze-quench EPR data from the reaction of the substrate complex of mixed-valent MIOX [MIOX(II/III)-MI] with limiting O2 in the presence of excess, saturating MI reveal the following cycle: (1) reacts rapidly with O2 to generate an intermediate (H) with a rhombic, g < 2 EPR spectrum; (2) a form of the enzyme with the same absorption features as MIOX(II/III) develops as H decays, suggesting that turnover has occurred; and (3) the starting MIOX(II/III)-MI complex is then quantitatively regenerated. This cycle is fast enough to account for the catalytic rate. The DG/O2 stoichiometry in the reaction, 0.8 ± 0.1, is similar to the theoretical value of 1, whereas significantly less product is formed in the corresponding reaction of the fully reduced enzyme with limiting O2. The DG/O2 yield in the latter reaction decreases as the enzyme concentration is increased, consistent with the hypothesis that initial conversion of the reduced enzyme to the MIOX(II/III)·MI complex and subsequent turnover by the mixed-valent form is responsible for the product in this case. The use of the mixed-valent, diiron(II/III) cluster by MIOX represents a significant departure from the mechanisms of other known diiron oxygenases, which all involve activation of O2 from the II/II manifold.

Original languageEnglish (US)
Pages (from-to)5402-5412
Number of pages11
JournalBiochemistry
Volume45
Issue number17
DOIs
StatePublished - May 2 2006

Fingerprint

Inositol Oxygenase
Glycols
Inositol
Chemical activation
Oxygen
Glucuronic Acid
Enzymes
Electron Spin Resonance Spectroscopy
Paramagnetic resonance
Stoichiometry
Oxygenases
Biochemistry
Substrates
Oxidation-Reduction
Cysteine
Spectrum Analysis
Iron

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Xing, Gang ; Barr, Eric W. ; Diao, Yinghui ; Hoffart, Lee M. ; Prabhu, Kumble Sandeep ; Arner, Ryan J. ; Reddy, C. Channa ; Krebs, Carsten ; Bollinger, Jr., Joseph M. / Oxygen activation by a mixed-valent, diiron(II/III) cluster in the glycol cleavage reaction catalyzed by myo-inositol oxygenase. In: Biochemistry. 2006 ; Vol. 45, No. 17. pp. 5402-5412.
@article{f37a0f85248f48739e28105196034f8f,
title = "Oxygen activation by a mixed-valent, diiron(II/III) cluster in the glycol cleavage reaction catalyzed by myo-inositol oxygenase",
abstract = "myo-Inositol oxygenase (MIOX) catalyzes the ring-cleaving, four-electron oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate (myo-inositol, MI) to D-glucuronate (DG). The preceding paper [Xing, G., Hoffart, L. M., Diao, Y., Prabhu, K. S., Arner, R. J., Reddy, C. C., Krebs, C., and Bollinger, J. M., Jr. (2006) Biochemistry 45, 5393-5401] demonstrates by M{\"o}ssbauer and electron paramagnetic resonance (EPR) spectroscopies that MIOX can contain a non-heme dinuclear iron cluster, which, in its mixed-valent (II/III) and fully oxidized (III/III) states, is perturbed by binding of MI in a manner consistent with direct coordination. In the study presented here, the redox form of the enzyme that activates O2 has been identified. L-Cysteine, which was previously reported to accelerate turnover, reduces the fully oxidized enzyme to the mixed-valent form, and O2, the cosubstrate, oxidizes the fully reduced form to the mixed-valent form with a stoichiometry of one per O 2, Both observations implicate the mixed-valent, diiron-(II/III) form of the enzyme as the active state. Stopped-flow absorption and freeze-quench EPR data from the reaction of the substrate complex of mixed-valent MIOX [MIOX(II/III)-MI] with limiting O2 in the presence of excess, saturating MI reveal the following cycle: (1) reacts rapidly with O2 to generate an intermediate (H) with a rhombic, g < 2 EPR spectrum; (2) a form of the enzyme with the same absorption features as MIOX(II/III) develops as H decays, suggesting that turnover has occurred; and (3) the starting MIOX(II/III)-MI complex is then quantitatively regenerated. This cycle is fast enough to account for the catalytic rate. The DG/O2 stoichiometry in the reaction, 0.8 ± 0.1, is similar to the theoretical value of 1, whereas significantly less product is formed in the corresponding reaction of the fully reduced enzyme with limiting O2. The DG/O2 yield in the latter reaction decreases as the enzyme concentration is increased, consistent with the hypothesis that initial conversion of the reduced enzyme to the MIOX(II/III)·MI complex and subsequent turnover by the mixed-valent form is responsible for the product in this case. The use of the mixed-valent, diiron(II/III) cluster by MIOX represents a significant departure from the mechanisms of other known diiron oxygenases, which all involve activation of O2 from the II/II manifold.",
author = "Gang Xing and Barr, {Eric W.} and Yinghui Diao and Hoffart, {Lee M.} and Prabhu, {Kumble Sandeep} and Arner, {Ryan J.} and Reddy, {C. Channa} and Carsten Krebs and {Bollinger, Jr.}, {Joseph M.}",
year = "2006",
month = "5",
day = "2",
doi = "10.1021/bi0526276",
language = "English (US)",
volume = "45",
pages = "5402--5412",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "17",

}

Oxygen activation by a mixed-valent, diiron(II/III) cluster in the glycol cleavage reaction catalyzed by myo-inositol oxygenase. / Xing, Gang; Barr, Eric W.; Diao, Yinghui; Hoffart, Lee M.; Prabhu, Kumble Sandeep; Arner, Ryan J.; Reddy, C. Channa; Krebs, Carsten; Bollinger, Jr., Joseph M.

In: Biochemistry, Vol. 45, No. 17, 02.05.2006, p. 5402-5412.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Oxygen activation by a mixed-valent, diiron(II/III) cluster in the glycol cleavage reaction catalyzed by myo-inositol oxygenase

AU - Xing, Gang

AU - Barr, Eric W.

AU - Diao, Yinghui

AU - Hoffart, Lee M.

AU - Prabhu, Kumble Sandeep

AU - Arner, Ryan J.

AU - Reddy, C. Channa

AU - Krebs, Carsten

AU - Bollinger, Jr., Joseph M.

PY - 2006/5/2

Y1 - 2006/5/2

N2 - myo-Inositol oxygenase (MIOX) catalyzes the ring-cleaving, four-electron oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate (myo-inositol, MI) to D-glucuronate (DG). The preceding paper [Xing, G., Hoffart, L. M., Diao, Y., Prabhu, K. S., Arner, R. J., Reddy, C. C., Krebs, C., and Bollinger, J. M., Jr. (2006) Biochemistry 45, 5393-5401] demonstrates by Mössbauer and electron paramagnetic resonance (EPR) spectroscopies that MIOX can contain a non-heme dinuclear iron cluster, which, in its mixed-valent (II/III) and fully oxidized (III/III) states, is perturbed by binding of MI in a manner consistent with direct coordination. In the study presented here, the redox form of the enzyme that activates O2 has been identified. L-Cysteine, which was previously reported to accelerate turnover, reduces the fully oxidized enzyme to the mixed-valent form, and O2, the cosubstrate, oxidizes the fully reduced form to the mixed-valent form with a stoichiometry of one per O 2, Both observations implicate the mixed-valent, diiron-(II/III) form of the enzyme as the active state. Stopped-flow absorption and freeze-quench EPR data from the reaction of the substrate complex of mixed-valent MIOX [MIOX(II/III)-MI] with limiting O2 in the presence of excess, saturating MI reveal the following cycle: (1) reacts rapidly with O2 to generate an intermediate (H) with a rhombic, g < 2 EPR spectrum; (2) a form of the enzyme with the same absorption features as MIOX(II/III) develops as H decays, suggesting that turnover has occurred; and (3) the starting MIOX(II/III)-MI complex is then quantitatively regenerated. This cycle is fast enough to account for the catalytic rate. The DG/O2 stoichiometry in the reaction, 0.8 ± 0.1, is similar to the theoretical value of 1, whereas significantly less product is formed in the corresponding reaction of the fully reduced enzyme with limiting O2. The DG/O2 yield in the latter reaction decreases as the enzyme concentration is increased, consistent with the hypothesis that initial conversion of the reduced enzyme to the MIOX(II/III)·MI complex and subsequent turnover by the mixed-valent form is responsible for the product in this case. The use of the mixed-valent, diiron(II/III) cluster by MIOX represents a significant departure from the mechanisms of other known diiron oxygenases, which all involve activation of O2 from the II/II manifold.

AB - myo-Inositol oxygenase (MIOX) catalyzes the ring-cleaving, four-electron oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate (myo-inositol, MI) to D-glucuronate (DG). The preceding paper [Xing, G., Hoffart, L. M., Diao, Y., Prabhu, K. S., Arner, R. J., Reddy, C. C., Krebs, C., and Bollinger, J. M., Jr. (2006) Biochemistry 45, 5393-5401] demonstrates by Mössbauer and electron paramagnetic resonance (EPR) spectroscopies that MIOX can contain a non-heme dinuclear iron cluster, which, in its mixed-valent (II/III) and fully oxidized (III/III) states, is perturbed by binding of MI in a manner consistent with direct coordination. In the study presented here, the redox form of the enzyme that activates O2 has been identified. L-Cysteine, which was previously reported to accelerate turnover, reduces the fully oxidized enzyme to the mixed-valent form, and O2, the cosubstrate, oxidizes the fully reduced form to the mixed-valent form with a stoichiometry of one per O 2, Both observations implicate the mixed-valent, diiron-(II/III) form of the enzyme as the active state. Stopped-flow absorption and freeze-quench EPR data from the reaction of the substrate complex of mixed-valent MIOX [MIOX(II/III)-MI] with limiting O2 in the presence of excess, saturating MI reveal the following cycle: (1) reacts rapidly with O2 to generate an intermediate (H) with a rhombic, g < 2 EPR spectrum; (2) a form of the enzyme with the same absorption features as MIOX(II/III) develops as H decays, suggesting that turnover has occurred; and (3) the starting MIOX(II/III)-MI complex is then quantitatively regenerated. This cycle is fast enough to account for the catalytic rate. The DG/O2 stoichiometry in the reaction, 0.8 ± 0.1, is similar to the theoretical value of 1, whereas significantly less product is formed in the corresponding reaction of the fully reduced enzyme with limiting O2. The DG/O2 yield in the latter reaction decreases as the enzyme concentration is increased, consistent with the hypothesis that initial conversion of the reduced enzyme to the MIOX(II/III)·MI complex and subsequent turnover by the mixed-valent form is responsible for the product in this case. The use of the mixed-valent, diiron(II/III) cluster by MIOX represents a significant departure from the mechanisms of other known diiron oxygenases, which all involve activation of O2 from the II/II manifold.

UR - http://www.scopus.com/inward/record.url?scp=33646347856&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33646347856&partnerID=8YFLogxK

U2 - 10.1021/bi0526276

DO - 10.1021/bi0526276

M3 - Article

VL - 45

SP - 5402

EP - 5412

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 17

ER -