Reversible Oxidative Modifications in Myoglobin and Functional Implications
Myoglobin (Mb), an oxygen-binding heme protein highly expressed in heart and skeletal muscle, has been shown to undergo oxidative modifications on both an inter- and intramolecular level when exposed to hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) in vitro. Here, we show th...
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| Format: | Article |
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MDPI AG
2020-06-01
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| Series: | Antioxidants |
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| Online Access: | https://www.mdpi.com/2076-3921/9/6/549 |
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| author | Mark H. Mannino Rishi S. Patel Amanda M. Eccardt Blythe E. Janowiak David C. Wood Fahu He Jonathan S. Fisher |
| author_facet | Mark H. Mannino Rishi S. Patel Amanda M. Eccardt Blythe E. Janowiak David C. Wood Fahu He Jonathan S. Fisher |
| author_sort | Mark H. Mannino |
| collection | DOAJ |
| description | Myoglobin (Mb), an oxygen-binding heme protein highly expressed in heart and skeletal muscle, has been shown to undergo oxidative modifications on both an inter- and intramolecular level when exposed to hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) in vitro. Here, we show that exposure to H<sub>2</sub>O<sub>2</sub> increases the peroxidase activity of Mb. Reaction of Mb with H<sub>2</sub>O<sub>2</sub> causes covalent binding of heme to the Mb protein (Mb-X), corresponding to an increase in peroxidase activity when ascorbic acid is the reducing co-substrate. Treatment of H<sub>2</sub>O<sub>2</sub>-reacted Mb with ascorbic acid reverses the Mb-X crosslink. Reaction with H<sub>2</sub>O<sub>2</sub> causes Mb to form dimers, trimers, and larger molecular weight Mb aggregates, and treatment with ascorbic acid regenerates Mb monomers. Reaction of Mb with H<sub>2</sub>O<sub>2</sub> causes formation of dityrosine crosslinks, though the labile nature of the crosslinks broken by treatment with ascorbic acid suggests that the reversible aggregation of Mb is mediated by crosslinks other than dityrosine. Disappearance of a peptide containing a tryptophan residue when Mb is treated with H<sub>2</sub>O<sub>2</sub> and the peptide’s reappearance after subsequent treatment with ascorbic acid suggest that tryptophan side chains might participate in the labile crosslinking. Taken together, these data suggest that while exposure to H<sub>2</sub>O<sub>2</sub> causes Mb-X formation, increases Mb peroxidase activity, and causes Mb aggregation, these oxidative modifications are reversible by treatment with ascorbic acid. A caveat is that future studies should demonstrate that these and other in vitro findings regarding properties of Mb have relevance in the intracellular milieu, especially in regard to actual concentrations of metMb, H<sub>2</sub>O<sub>2</sub>, and ascorbate that would be found in vivo. |
| first_indexed | 2024-03-10T18:55:22Z |
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| id | doaj.art-79efca5a14f14b159e54902ee85d0482 |
| institution | Directory Open Access Journal |
| issn | 2076-3921 |
| language | English |
| last_indexed | 2024-03-10T18:55:22Z |
| publishDate | 2020-06-01 |
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| series | Antioxidants |
| spelling | doaj.art-79efca5a14f14b159e54902ee85d04822023-11-20T04:48:40ZengMDPI AGAntioxidants2076-39212020-06-019654910.3390/antiox9060549Reversible Oxidative Modifications in Myoglobin and Functional ImplicationsMark H. Mannino0Rishi S. Patel1Amanda M. Eccardt2Blythe E. Janowiak3David C. Wood4Fahu He5Jonathan S. Fisher6Department of Biology, Saint Louis University, St. Louis, MO 63103, USADepartment of Biology, Saint Louis University, St. Louis, MO 63103, USADepartment of Biology, Saint Louis University, St. Louis, MO 63103, USADepartment of Biology, Saint Louis University, St. Louis, MO 63103, USADepartment of Biochemistry and Molecular Biology, Saint Louis University, St. Louis, MO 63104, USADepartment of Chemistry, Saint Louis University, St. Louis, MO 63103, USADepartment of Biology, Saint Louis University, St. Louis, MO 63103, USAMyoglobin (Mb), an oxygen-binding heme protein highly expressed in heart and skeletal muscle, has been shown to undergo oxidative modifications on both an inter- and intramolecular level when exposed to hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) in vitro. Here, we show that exposure to H<sub>2</sub>O<sub>2</sub> increases the peroxidase activity of Mb. Reaction of Mb with H<sub>2</sub>O<sub>2</sub> causes covalent binding of heme to the Mb protein (Mb-X), corresponding to an increase in peroxidase activity when ascorbic acid is the reducing co-substrate. Treatment of H<sub>2</sub>O<sub>2</sub>-reacted Mb with ascorbic acid reverses the Mb-X crosslink. Reaction with H<sub>2</sub>O<sub>2</sub> causes Mb to form dimers, trimers, and larger molecular weight Mb aggregates, and treatment with ascorbic acid regenerates Mb monomers. Reaction of Mb with H<sub>2</sub>O<sub>2</sub> causes formation of dityrosine crosslinks, though the labile nature of the crosslinks broken by treatment with ascorbic acid suggests that the reversible aggregation of Mb is mediated by crosslinks other than dityrosine. Disappearance of a peptide containing a tryptophan residue when Mb is treated with H<sub>2</sub>O<sub>2</sub> and the peptide’s reappearance after subsequent treatment with ascorbic acid suggest that tryptophan side chains might participate in the labile crosslinking. Taken together, these data suggest that while exposure to H<sub>2</sub>O<sub>2</sub> causes Mb-X formation, increases Mb peroxidase activity, and causes Mb aggregation, these oxidative modifications are reversible by treatment with ascorbic acid. A caveat is that future studies should demonstrate that these and other in vitro findings regarding properties of Mb have relevance in the intracellular milieu, especially in regard to actual concentrations of metMb, H<sub>2</sub>O<sub>2</sub>, and ascorbate that would be found in vivo.https://www.mdpi.com/2076-3921/9/6/549myoglobindityrosineditryptophanperoxidaseprotein aggregation |
| spellingShingle | Mark H. Mannino Rishi S. Patel Amanda M. Eccardt Blythe E. Janowiak David C. Wood Fahu He Jonathan S. Fisher Reversible Oxidative Modifications in Myoglobin and Functional Implications Antioxidants myoglobin dityrosine ditryptophan peroxidase protein aggregation |
| title | Reversible Oxidative Modifications in Myoglobin and Functional Implications |
| title_full | Reversible Oxidative Modifications in Myoglobin and Functional Implications |
| title_fullStr | Reversible Oxidative Modifications in Myoglobin and Functional Implications |
| title_full_unstemmed | Reversible Oxidative Modifications in Myoglobin and Functional Implications |
| title_short | Reversible Oxidative Modifications in Myoglobin and Functional Implications |
| title_sort | reversible oxidative modifications in myoglobin and functional implications |
| topic | myoglobin dityrosine ditryptophan peroxidase protein aggregation |
| url | https://www.mdpi.com/2076-3921/9/6/549 |
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