Nitric Oxide pp 281-297 | Cite as

Thioredoxin-Dependent Decomposition of Protein S-Nitrosothiols

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1747)

Abstract

The addition of nitric oxide to cysteine moieties of proteins results in the formation of S-nitrosothiols (SNO) that have emerged as important posttranslational signaling cues in a wide variety of eukaryotic processes. While formation of protein-SNO is largely nonenzymatic, the conserved family of Thioredoxin (TRX) enzymes are capable of selectively reducing protein-SNO. Consequently, TRX enzymes are thought to provide reversibility and specificity to protein-SNO signaling networks. Here, we describe an in vitro methodology based on enzymatic oxidoreductase and biotin-switch techniques, allowing for the detection of protein-SNO targets of TRX enzymes. We show that this methodology identifies both global and specific protein-SNO targets of TRX in plant cell extracts.

Key words

Biotin switch technique S-nitrosylation Denitrosylation Thioredoxin Protein-SNO reductase Nitric oxide 

Notes

Acknowledgments

This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement No 678511), a Royal Society University Research Fellowship (UF090321), and a Wellcome Trust-University of Edinburgh Institutional Strategic Support Fund (ISSF).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Institute of Molecular Plant Sciences, School of Biological SciencesUniversity of EdinburghEdinburghUK

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