Stoichiometric production of hydrogen peroxide and parallel formation of ferric multimers through decay of the diferric-peroxo complex, the first detectable intermediate in ferritin mineralization

Guy N L Jameson, Weili Jin, Carsten Krebs, Alice S. Perreira, Pedro Tavares, Xiaofeng Liu, Elizabeth C. Theil, Boi Hanh Huynh

Research output: Contribution to journalArticle

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Abstract

The catalytic step that initiates formation of the ferric oxy-hydroxide mineral core in the central cavity of H-type ferritin involves rapid oxidation of ferrous ion by molecular oxygen (ferroxidase reaction) at a binuclear site (ferroxidase site) found in each of the 24 subunits. Previous investigators have shown that the first detectable reaction intermediate of the ferroxidase reaction is a diferric-peroxo intermediate, Fperoxo, formed within 25 ms, which then leads to the release of H2O2 and formation of ferric mineral precursors. The stoichiometric relationship between Fperoxo, H2O2, and ferric mineral precursors, crucial to defining the reaction pathway and mechanism, has now been determined. To this end, a horseradish peroxidase-catalyzed spectrophotometric method was used as an assay for H2O2. By rapidly mixing apo M ferritin from frog, Fe2+, and O2 and allowing the reaction to proceed for 70 ms when Fperoxo has reached its maximum accumulation, followed by spraying the reaction mixture into the H2O2 assay solution, we were able to quantitatively determine the amount of H2O2 produced during the decay of Fperoxo. The correlation between the amount of H2O2 released with the amount of Fperoxo accumulated at 70 ms determined by Mössbauer spectroscopy showed that Fperoxo decays into H2O2 with a stoichiometry of 1 Fperoxo:H2O2. When the decay of Fperoxo was monitored by rapid freeze - quench Mössbauer spectroscopy, multiple diferric μ-oxo/μ-hydroxo complexes and small polynuclear ferric clusters were found to form at rate constants identical to the decay rate of Fperoxo. This observed parallel formation of multiple products (H2O2, diferric complexes, and small polynuclear clusters) from the decay of a single precursor (Fperoxo) provides useful mechanistic insights into ferritin mineralization and demonstrates a flexible ferroxidase site.

Original languageEnglish (US)
Pages (from-to)13435-13443
Number of pages9
JournalBiochemistry
Volume41
Issue number45
DOIs
StatePublished - Nov 12 2002

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Ceruloplasmin
Ferritins
Hydrogen Peroxide
Apoferritins
Minerals
Assays
Spectrum Analysis
Spectroscopy
Reaction intermediates
Molecular oxygen
Horseradish Peroxidase
Spraying
Stoichiometry
Anura
Rate constants
Research Personnel
Ions
Oxygen
Oxidation

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Jameson, Guy N L ; Jin, Weili ; Krebs, Carsten ; Perreira, Alice S. ; Tavares, Pedro ; Liu, Xiaofeng ; Theil, Elizabeth C. ; Huynh, Boi Hanh. / Stoichiometric production of hydrogen peroxide and parallel formation of ferric multimers through decay of the diferric-peroxo complex, the first detectable intermediate in ferritin mineralization. In: Biochemistry. 2002 ; Vol. 41, No. 45. pp. 13435-13443.
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abstract = "The catalytic step that initiates formation of the ferric oxy-hydroxide mineral core in the central cavity of H-type ferritin involves rapid oxidation of ferrous ion by molecular oxygen (ferroxidase reaction) at a binuclear site (ferroxidase site) found in each of the 24 subunits. Previous investigators have shown that the first detectable reaction intermediate of the ferroxidase reaction is a diferric-peroxo intermediate, Fperoxo, formed within 25 ms, which then leads to the release of H2O2 and formation of ferric mineral precursors. The stoichiometric relationship between Fperoxo, H2O2, and ferric mineral precursors, crucial to defining the reaction pathway and mechanism, has now been determined. To this end, a horseradish peroxidase-catalyzed spectrophotometric method was used as an assay for H2O2. By rapidly mixing apo M ferritin from frog, Fe2+, and O2 and allowing the reaction to proceed for 70 ms when Fperoxo has reached its maximum accumulation, followed by spraying the reaction mixture into the H2O2 assay solution, we were able to quantitatively determine the amount of H2O2 produced during the decay of Fperoxo. The correlation between the amount of H2O2 released with the amount of Fperoxo accumulated at 70 ms determined by M{\"o}ssbauer spectroscopy showed that Fperoxo decays into H2O2 with a stoichiometry of 1 Fperoxo:H2O2. When the decay of Fperoxo was monitored by rapid freeze - quench M{\"o}ssbauer spectroscopy, multiple diferric μ-oxo/μ-hydroxo complexes and small polynuclear ferric clusters were found to form at rate constants identical to the decay rate of Fperoxo. This observed parallel formation of multiple products (H2O2, diferric complexes, and small polynuclear clusters) from the decay of a single precursor (Fperoxo) provides useful mechanistic insights into ferritin mineralization and demonstrates a flexible ferroxidase site.",
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Stoichiometric production of hydrogen peroxide and parallel formation of ferric multimers through decay of the diferric-peroxo complex, the first detectable intermediate in ferritin mineralization. / Jameson, Guy N L; Jin, Weili; Krebs, Carsten; Perreira, Alice S.; Tavares, Pedro; Liu, Xiaofeng; Theil, Elizabeth C.; Huynh, Boi Hanh.

In: Biochemistry, Vol. 41, No. 45, 12.11.2002, p. 13435-13443.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Stoichiometric production of hydrogen peroxide and parallel formation of ferric multimers through decay of the diferric-peroxo complex, the first detectable intermediate in ferritin mineralization

AU - Jameson, Guy N L

AU - Jin, Weili

AU - Krebs, Carsten

AU - Perreira, Alice S.

AU - Tavares, Pedro

AU - Liu, Xiaofeng

AU - Theil, Elizabeth C.

AU - Huynh, Boi Hanh

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Y1 - 2002/11/12

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