Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34
Human Mn-containing superoxide dismutase (hMnSOD) is a mitochondrial enzyme that metabolizes superoxide radical (O2 •-). O2 •- reacts at diffusional rates with nitric oxide to yield a potent nitrating species, peroxynitrite anion (ONOO-). MnSOD is nitrated and inactivated in vivo, with active site T...
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2016
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paper:paper_00062960_v55_n24_p3403_Demicheli2023-06-08T14:30:49Z Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 Jara, Gabriel Ernesto Martí, Marcelo Adrián Amino acids Enzymes Fluorescence Manganese Molecular modeling Nitric oxide Oxygen Quantum theory Rate constants Reaction intermediates Atomic-level resolution Manganese superoxide dismutase Mitochondrial enzymes Quantum mechanics/molecular mechanics Second-order rate constants Selective modification Super oxide dismutase Time-resolved fluorescence Nitration 3 nitrotyrosine boronic acid derivative coumarin coumarin boronic acid dithionite manganese superoxide dismutase monomer peroxynitrite sodium azide transition element tyrosine tyrosine 34 unclassified drug 7-hydroxycoumarin nitric acid derivative nitric oxide peroxynitrous acid superoxide dismutase tyrosine umbelliferone derivative Article enzyme activity enzyme inactivation fluorescence analysis human isoelectric point kinetics molecular mechanics nitration oxidation reduction potential oxidation reduction reaction priority journal proteomics quantum mechanics rate constant two dimensional electrophoresis enzyme active site metabolism molecular model Western blotting Blotting, Western Catalytic Domain Humans Kinetics Models, Molecular Nitrates Nitric Oxide Oxidation-Reduction Peroxynitrous Acid Proteomics Superoxide Dismutase Tyrosine Umbelliferones Human Mn-containing superoxide dismutase (hMnSOD) is a mitochondrial enzyme that metabolizes superoxide radical (O2 •-). O2 •- reacts at diffusional rates with nitric oxide to yield a potent nitrating species, peroxynitrite anion (ONOO-). MnSOD is nitrated and inactivated in vivo, with active site Tyr34 as the key oxidatively modified residue. We previously reported a k of ?1.0 × 105 M-1 s-1 for the reaction of hMnSOD with ONOO- by direct stopped-flow spectroscopy and the critical role of Mn in the nitration process. In this study, we further established the mechanism of the reaction of hMnSOD with ONOO-, including the necessary re-examination of the second-order rate constant by an independent method and the delineation of the microscopic steps that lead to the regio-specific nitration of Tyr34. The redetermination of k was performed by competition kinetics utilizing coumarin boronic acid, which reacts with ONOO- at a rate of ?1 × 106 M-1 s-1 to yield the fluorescence product, 7-hydroxycoumarin. Time-resolved fluorescence studies in the presence of increasing concentrations of hMnSOD provided a k of ?1.0 × 105 M-1 s-1, fully consistent with the direct method. Proteomic analysis indicated that ONOO-, but not other nitrating agents, mediates the selective modification of active site Tyr34. Hybrid quantum-classical (quantum mechanics/molecular mechanics) simulations supported a series of steps that involve the initial reaction of ONOO- with MnIII to yield MnIV and intermediates that ultimately culminate in 3-nitroTyr34. The data reported herein provide a kinetic and mechanistic basis for rationalizing how MnSOD constitutes an intramitochondrial target for ONOO- and the microscopic events, with atomic level resolution, that lead to selective and efficient nitration of critical Tyr34. © 2016 American Chemical Society. Fil:Jara, G.E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Martí, M.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. 2016 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00062960_v55_n24_p3403_Demicheli http://hdl.handle.net/20.500.12110/paper_00062960_v55_n24_p3403_Demicheli |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Amino acids Enzymes Fluorescence Manganese Molecular modeling Nitric oxide Oxygen Quantum theory Rate constants Reaction intermediates Atomic-level resolution Manganese superoxide dismutase Mitochondrial enzymes Quantum mechanics/molecular mechanics Second-order rate constants Selective modification Super oxide dismutase Time-resolved fluorescence Nitration 3 nitrotyrosine boronic acid derivative coumarin coumarin boronic acid dithionite manganese superoxide dismutase monomer peroxynitrite sodium azide transition element tyrosine tyrosine 34 unclassified drug 7-hydroxycoumarin nitric acid derivative nitric oxide peroxynitrous acid superoxide dismutase tyrosine umbelliferone derivative Article enzyme activity enzyme inactivation fluorescence analysis human isoelectric point kinetics molecular mechanics nitration oxidation reduction potential oxidation reduction reaction priority journal proteomics quantum mechanics rate constant two dimensional electrophoresis enzyme active site metabolism molecular model Western blotting Blotting, Western Catalytic Domain Humans Kinetics Models, Molecular Nitrates Nitric Oxide Oxidation-Reduction Peroxynitrous Acid Proteomics Superoxide Dismutase Tyrosine Umbelliferones |
| spellingShingle |
Amino acids Enzymes Fluorescence Manganese Molecular modeling Nitric oxide Oxygen Quantum theory Rate constants Reaction intermediates Atomic-level resolution Manganese superoxide dismutase Mitochondrial enzymes Quantum mechanics/molecular mechanics Second-order rate constants Selective modification Super oxide dismutase Time-resolved fluorescence Nitration 3 nitrotyrosine boronic acid derivative coumarin coumarin boronic acid dithionite manganese superoxide dismutase monomer peroxynitrite sodium azide transition element tyrosine tyrosine 34 unclassified drug 7-hydroxycoumarin nitric acid derivative nitric oxide peroxynitrous acid superoxide dismutase tyrosine umbelliferone derivative Article enzyme activity enzyme inactivation fluorescence analysis human isoelectric point kinetics molecular mechanics nitration oxidation reduction potential oxidation reduction reaction priority journal proteomics quantum mechanics rate constant two dimensional electrophoresis enzyme active site metabolism molecular model Western blotting Blotting, Western Catalytic Domain Humans Kinetics Models, Molecular Nitrates Nitric Oxide Oxidation-Reduction Peroxynitrous Acid Proteomics Superoxide Dismutase Tyrosine Umbelliferones Jara, Gabriel Ernesto Martí, Marcelo Adrián Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 |
| topic_facet |
Amino acids Enzymes Fluorescence Manganese Molecular modeling Nitric oxide Oxygen Quantum theory Rate constants Reaction intermediates Atomic-level resolution Manganese superoxide dismutase Mitochondrial enzymes Quantum mechanics/molecular mechanics Second-order rate constants Selective modification Super oxide dismutase Time-resolved fluorescence Nitration 3 nitrotyrosine boronic acid derivative coumarin coumarin boronic acid dithionite manganese superoxide dismutase monomer peroxynitrite sodium azide transition element tyrosine tyrosine 34 unclassified drug 7-hydroxycoumarin nitric acid derivative nitric oxide peroxynitrous acid superoxide dismutase tyrosine umbelliferone derivative Article enzyme activity enzyme inactivation fluorescence analysis human isoelectric point kinetics molecular mechanics nitration oxidation reduction potential oxidation reduction reaction priority journal proteomics quantum mechanics rate constant two dimensional electrophoresis enzyme active site metabolism molecular model Western blotting Blotting, Western Catalytic Domain Humans Kinetics Models, Molecular Nitrates Nitric Oxide Oxidation-Reduction Peroxynitrous Acid Proteomics Superoxide Dismutase Tyrosine Umbelliferones |
| description |
Human Mn-containing superoxide dismutase (hMnSOD) is a mitochondrial enzyme that metabolizes superoxide radical (O2 •-). O2 •- reacts at diffusional rates with nitric oxide to yield a potent nitrating species, peroxynitrite anion (ONOO-). MnSOD is nitrated and inactivated in vivo, with active site Tyr34 as the key oxidatively modified residue. We previously reported a k of ?1.0 × 105 M-1 s-1 for the reaction of hMnSOD with ONOO- by direct stopped-flow spectroscopy and the critical role of Mn in the nitration process. In this study, we further established the mechanism of the reaction of hMnSOD with ONOO-, including the necessary re-examination of the second-order rate constant by an independent method and the delineation of the microscopic steps that lead to the regio-specific nitration of Tyr34. The redetermination of k was performed by competition kinetics utilizing coumarin boronic acid, which reacts with ONOO- at a rate of ?1 × 106 M-1 s-1 to yield the fluorescence product, 7-hydroxycoumarin. Time-resolved fluorescence studies in the presence of increasing concentrations of hMnSOD provided a k of ?1.0 × 105 M-1 s-1, fully consistent with the direct method. Proteomic analysis indicated that ONOO-, but not other nitrating agents, mediates the selective modification of active site Tyr34. Hybrid quantum-classical (quantum mechanics/molecular mechanics) simulations supported a series of steps that involve the initial reaction of ONOO- with MnIII to yield MnIV and intermediates that ultimately culminate in 3-nitroTyr34. The data reported herein provide a kinetic and mechanistic basis for rationalizing how MnSOD constitutes an intramitochondrial target for ONOO- and the microscopic events, with atomic level resolution, that lead to selective and efficient nitration of critical Tyr34. © 2016 American Chemical Society. |
| author |
Jara, Gabriel Ernesto Martí, Marcelo Adrián |
| author_facet |
Jara, Gabriel Ernesto Martí, Marcelo Adrián |
| author_sort |
Jara, Gabriel Ernesto |
| title |
Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 |
| title_short |
Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 |
| title_full |
Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 |
| title_fullStr |
Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 |
| title_full_unstemmed |
Mechanism of the Reaction of Human Manganese Superoxide Dismutase with Peroxynitrite: Nitration of Critical Tyrosine 34 |
| title_sort |
mechanism of the reaction of human manganese superoxide dismutase with peroxynitrite: nitration of critical tyrosine 34 |
| publishDate |
2016 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00062960_v55_n24_p3403_Demicheli http://hdl.handle.net/20.500.12110/paper_00062960_v55_n24_p3403_Demicheli |
| work_keys_str_mv |
AT jaragabrielernesto mechanismofthereactionofhumanmanganesesuperoxidedismutasewithperoxynitritenitrationofcriticaltyrosine34 AT martimarceloadrian mechanismofthereactionofhumanmanganesesuperoxidedismutasewithperoxynitritenitrationofcriticaltyrosine34 |
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1768544892658647040 |