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Proteomic analysis of peroxynitrite-induced protein nitration in isolated beef heart mitochondria
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SYSNO ASEP 0489848 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Proteomic analysis of peroxynitrite-induced protein nitration in isolated beef heart mitochondria Author(s) Kohutiar, Matej (FGU-C)
Eckhardt, Adam (FGU-C) RID, ORCID
Mikšík, Ivan (FGU-C) RID, ORCID, SAI
Šantorová, J. (CZ)
Wilhelm, Jiří (FGU-C) RIDSource Title Physiological Research. - : Fyziologický ústav AV ČR, v. v. i. - ISSN 0862-8408
Roč. 67, č. 2 (2018), s. 239-250Number of pages 12 s. Language eng - English Country CZ - Czech Republic Keywords mitochondrial proteome ; peroxynitrite ; 3-nitrotyrosine ; 2D electrophoresis ; mass spectrometry Subject RIV CE - Biochemistry OECD category Biochemistry and molecular biology R&D Projects GAP303/11/0298 GA ČR - Czech Science Foundation (CSF) GA15-01948S GA ČR - Czech Science Foundation (CSF) Institutional support FGU-C - RVO:67985823 UT WOS 000431452400010 EID SCOPUS 85046813042 DOI 10.33549/physiolres.933608 Annotation Mitochondria are exposed to reactive nitrogen species under physiological conditions and even more under several pathologic states. In order to reveal the mechanism of these processes we studied the effects of peroxynitrite on isolated beef heart mitochondria in vitro. Peroxynitrite has the potential to nitrate protein tyrosine moieties, break the peptide bond, and eventually release the membrane proteins into the solution. All these effects were found in our experiments. Mitochondrial proteins were resolved by 2D electrophoresis and the protein nitration was detected by immunochemical methods and by nano LC-MS/MS. Mass spectrometry confirmed nitration of ATP synthase subunit beta, pyruvate dehydrogenase E1 component subunit beta, citrate synthase and acetyl-CoA acetyltransferase. Immunoblot detection using chemiluminiscence showed possible nitration of other proteins such as cytochrome b-c1 complex subunit 1, NADH dehydrogenase [ubiquinone] iron-sulfur protein 2, elongation factor Tu, NADH dehydrogenase [ubiquinone] flavoprotein 2, heat shock protein beta-1 and NADH dehydrogenase [ubiquinone] iron-sulfur protein 8. ATP synthase beta subunit was nitrated both in membrane and in fraction prepared by osmotic lysis. The high sensitivity of proteins to nitration by peroxynitrite is of potential biological importance, as these enzymes are involved in various pathways associated with energy production in the heart. Workplace Institute of Physiology Contact Lucie Trajhanová, lucie.trajhanova@fgu.cas.cz, Tel.: 241 062 400 Year of Publishing 2019
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