Enrichment and Detection of Tyrosine‐Nitrated Proteins

Abello, Nicolas; Wisastra, Rosalina; Bischoff, Rainer

doi:10.1002/0471140864.ps1413s69

Cited by 14 publications

(13 citation statements)

References 16 publications

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“…MS-based analyses of tyrosine-nitrated peptides include several different separation and enrichment procedures (see for instance Lee et al, 2009;Draguşanu et al, 2010;Draguşanu, Petre, & Przybylski, 2011;Dremina et al, 2011;Prokai-Tatrai, Guo, & Prokai, 2011). Some of these procedures have been reviewed recently (Nuriel et al, 2008;Abello et al, 2009;Dekker et al, 2012). For a brief summary see Table 6.…”

Section: Mass Spectrometric Analysis Of 3-nitrotyrosine In Peptimentioning

confidence: 99%

Mass spectrometry and 3-nitrotyrosine: Strategies, controversies, and our current perspective

Tsikas

Duncan

2013

Mass Spec Rev

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Reactive-nitrogen species (RNS) such as peroxynitrite (ONOO(-)), that is, the reaction product of nitric oxide ((•)NO) and superoxide (O2(-•)), nitryl chloride (NO2Cl) and (•)NO2 react with the activated aromatic ring of tyrosine to form 3-nitrotyrosine. This modification, which has been known for more than a century, occurs to both the free form of the amino acid (i.e., soluble/free tyrosine) and to tyrosine residues covalently bound within the backbone of peptides and proteins. Nitration of tyrosine is thought to be of biological significance and has been linked to health and disease, but determining its role has proved challenging. Several key questions have been the focus of much of the research activity: (a) to what extent is free/soluble tyrosine nitrated in biological tissues and fluids, and (b) are there specific site(s) of nitration within peptides/proteins and to what extent (i.e., stoichiometry) does this modification occur? These issues have been addressed in a wide range of sample types (e.g., blood, urine, CSF, exhaled breath condensate and various tissues) and a diverse array of physiological/pathophysiological scenarios. The accurate determination of nitrated tyrosine is, however, a stumbling block. Despite extensive study, the extent to which nitration occurs in vivo, the specificity of the nitration reaction, and its importance in health and disease, remain unclear. In this review, we highlight the analytical challenges and discuss the approaches adopted to address them. Mass spectrometry, in combination with either gas chromatography (GC-MS, GC-MS/MS) or liquid chromatography (LC-MS/MS), has played the central role in the analysis of 3-nitrotyrosine and tyrosine-nitrated biological macromolecules. We discuss its unique attributes and highlight the role of stable-isotope labeled 3-nitrotyrosine analogs in both accurate quantification, and in helping to define the biological relevance of tyrosine nitration. We show that the application of sophisticated mass spectrometric techniques is advantageous if not essential, but that this alone is by no means a guarantee of accurate findings. We discuss the important analytical challenges in quantifying 3-nitrotyrosine, possible workarounds, and we attempt to make sense of the disparate findings that have been reported so far.

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Section: Mass Spectrometric Analysis Of 3-nitrotyrosine In Peptimentioning

confidence: 99%

Mass spectrometry and 3-nitrotyrosine: Strategies, controversies, and our current perspective

Tsikas

Duncan

2013

Mass Spec Rev

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“…Thus, the combination of proteomic techniques (two-dimensional polyacryamide gel electrophoresis) followed by immunoblotting or immunoprecipitation, gel tryptic digestion and mass spectrometry have become powerful tools to develop nitroproteome studies in higher plants. Although there might be some technical problems like those due to the unspecific recognition of the antibody used against nitrotyrosine, the low number of nitrotyrosines in a given protein, the low abundance of the nitrotyrosine-containing proteins, and that the isolated protein occasionally does not match in the protein database, it must be pointed out that these approaches have allowed establishing a basic background of knowledge in this research area (Dekker et al, 2012). In any case, after a specific protein has been identified as a putative nitration target, a necessary additional step is to identify the nitration site(s) within the protein quaternary structure by MALDI-TOF MS and LC-MS/MS (Ytterberg and Jensen, 2010).…”

Section: In Vivo Detection Of Protein Targets Of Tyrosine Nitration Imentioning

confidence: 99%

Protein tyrosine nitration in higher plants grown under natural and stress conditions

Corpas¹,

Palma²,

Rı́o³

et al. 2013

Front. Plant Sci.

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Protein tyrosine nitration is a post-translational modification (PTM) mediated by reactive nitrogen species (RNS) that is linked to nitro-oxidative damages in plant cells. During the last decade, the identification of proteins undergoing this PTM under adverse environmental conditions has increased. However, there is also a basal endogenous nitration which seems to have a regulatory function. The technological advances in proteome analysis have allowed identifying these modified proteins and have shown that the number and identity of the nitrated proteins change among plant species, analysed organs and growing/culture conditions. In this work, the current knowledge of protein tyrosine nitration in higher plants under different situations is reviewed.

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“…Currently, an enrichment technique followed by MS is the most powerful approach for detecting nitropeptides. Enrichment procedures include use of either 3-NT specific antibodies or chemical derivation that reduces a nitro group to an amine group (17)(18)(19). Most previous studies aimed at methodology development and used in vitro overnitrated samples to profile artificially nitrated instead of endogenously nitrated proteins.…”

mentioning

confidence: 99%

Myeloid-derived suppressor cells inhibit T cell activation through nitrating LCK in mouse cancers

Feng

Cheng

Zhang

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

118

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Potent immunosuppressive mechanisms within the tumor microenvironment contribute to the resistance of aggressive human cancers to immune checkpoint blockade (ICB) therapy. One of the main mechanisms for myeloid-derived suppressor cells (MDSCs) to induce T cell tolerance is through secretion of reactive nitrogen species (RNS), which nitrates tyrosine residues in proteins involved in T cell function. However, so far very few nitrated proteins have been identified. Here, using a transgenic mouse model of prostate cancer and a syngeneic cell line model of lung cancer, we applied a nitroproteomic approach based on chemical derivation of 3-nitrotyrosine and identified that lymphocyte-specific protein tyrosine kinase (LCK), an initiating tyrosine kinase in the T cell receptor signaling cascade, is nitrated at Tyr394 by MDSCs. LCK nitration inhibits T cell activation, leading to reduced interleukin 2 (IL2) production and proliferation. In human T cells with defective endogenous LCK, wild type, but not nitrated LCK, rescues IL2 production. In the mouse model of castration-resistant prostate cancer (CRPC) by prostate-specific deletion of ,, and , CRPC is resistant to an ICB therapy composed of antiprogrammed cell death 1 (PD1) and anticytotoxic-T lymphocyte-associated protein 4 (CTLA4) antibodies. However, we showed that ICB elicits strong anti-CRPC efficacy when combined with an RNS neutralizing agent. Together, these data identify a previously unknown mechanism of T cell inactivation by MDSC-induced protein nitration and illuminate a clinical path hypothesis for combining ICB with RNS-reducing agents in the treatment of CRPC.

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Enrichment and Detection of Tyrosine‐Nitrated Proteins

Cited by 14 publications

References 16 publications

Mass spectrometry and 3-nitrotyrosine: Strategies, controversies, and our current perspective

Mass spectrometry and 3-nitrotyrosine: Strategies, controversies, and our current perspective

Protein tyrosine nitration in higher plants grown under natural and stress conditions

Myeloid-derived suppressor cells inhibit T cell activation through nitrating LCK in mouse cancers

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