A tale of two controversies. Defining both the role of peroxidases in nitrotyrosine formation in vivo using eosinophil peroxidase and myeloperoxidase-deficient mice, and the nature of peroxidase-generated reactive nitrogen species

Marie Luise Brennan, Weijia Wu, Xiaoming Fu, Zhongzhu Shen, Wei Song, Heather Frost, Caryn Vadseth, Laura Narine, Elizabeth Lenkiewicz, Michael T. Borchers, Aldons J. Lusis, James J. Lee, Nancy A. Lee, Husam M. Abu-Soud, Harry Ischiropoulos, Stanley L. Hazen

Research output: Contribution to journalArticle

430 Scopus citations

Abstract

Nitrotyrosine is widely used as a marker of post-translational modification by the nitric oxide (.NO, nitrogen monoxide)-derived oxidant peroxynitrite (ONOO-). However, since the discovery that myeloperoxidase (MPO) and eosinophil peroxidase (EPO) can generate nitrotyrosine via oxidation of nitrite (NO2-), several questions have arisen. First, the relative contribution of peroxidases to nitrotyrosine formation in vivo is unknown. Further, although evidence suggests that the one-electron oxidation product, nitrogen dioxide (.NO2), is the primary species formed, neither a direct demonstration that peroxidases form this gas nor studies designed to test for the possible concomitant formation of the two-electron oxidation product, ONOO-, have been reported. Using multiple distinct models of acute inflammation with EPO- and MPO-knockout mice, we now demonstrate that leukocyte peroxidases participate in nitrotyrosine formation in vivo. In some models, MPO and EPO played a dominant role, accounting for the majority of nitrotyrosine formed. However, in other leukocyte-rich acute inflammatory models, no contribution for either MPO or EPO to nitrotyrosine formation could be demonstrated. Head-space gas analysis of heliumswept reaction mixtures provides direct evidence that leukocyte peroxidases catalytically generate .NO2 formation using H2O2 and NO2- as substrates. However, formation of an additional oxidant was suggested since both enzymes promote NO2--dependent hydroxylation of targets under acidic conditions, a chemical reactivity shared with ONOO- but not .NO2. Collectively, our results demonstrate that: 1) MPO and EPO contribute to tyrosine nitration in vivo; 2) the major reactive nitrogen species formed by leukocyte peroxidase-catalyzed oxidation of NO2- is the one-electron oxidation product, ̇NO2; 3) as a minor reaction, peroxidases may also catalyze the two-electron oxidation of NO2-, producing a ONOO--like product. We speculate that the latter reaction generates a labile Fe-ONOO complex, which may be released following protonation under acidic conditions such as might exist at sites of inflammation.

Original languageEnglish (US)
Pages (from-to)17415-17427
Number of pages13
JournalJournal of Biological Chemistry
Volume277
Issue number20
DOIs
StatePublished - May 17 2002

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Brennan, M. L., Wu, W., Fu, X., Shen, Z., Song, W., Frost, H., Vadseth, C., Narine, L., Lenkiewicz, E., Borchers, M. T., Lusis, A. J., Lee, J. J., Lee, N. A., Abu-Soud, H. M., Ischiropoulos, H., & Hazen, S. L. (2002). A tale of two controversies. Defining both the role of peroxidases in nitrotyrosine formation in vivo using eosinophil peroxidase and myeloperoxidase-deficient mice, and the nature of peroxidase-generated reactive nitrogen species. Journal of Biological Chemistry, 277(20), 17415-17427. https://doi.org/10.1074/jbc.M112400200