Improved Tag-Switch Method Reveals that Thioredoxin Acts as Depersulfidase and Controls the Intracellular Levels of Protein Persulfidation
András Szijártó, I
Royal Society of Chemistry
This is the final version of the article. It first appeared from Royal Society of Chemistry via http://dx.doi.org/10.1039/C5SC04818D.
Hydrogen sulfide (H2S) has emerged as a signalling molecule capable of regulating several important physiological functions such as blood pressure, neurotransmission and inflammation. The mechanisms behind these effects are still largely elusive and oxidative posttranslational modification of cysteine residues (protein persulfidation or S-sulfhydration) has been proposed as the main pathway for H2S-induced biological and pharmacological effects. As a signalling mechanism, persulfidation has to be controlled. Using an improved tag-switch assay for persulfide detection we show here that protein persulfide levels are controlled by the thioredoxin system. Recombinant thioredoxin showed an almost 10-fold higher reactivity towards cysteine persulfide than towards cystine and readily cleaved protein persulfides as well. This reaction resulted in H2S release suggesting that thioredoxin could be an important regulator of H2S levels from persulfide pools. Inhibition of the thioredoxin system caused an increase in intracellular persulfides, highlighting thioredoxin as a major protein depersulfidase that controls H2S signalling. Finally, using plasma from HIV-1 patients that have higher circulatory levels of thioredoxin, we could prove depersulfidase role in vivo.
This research was supported by an intramural grant from Friedrich-Alexander University within the Emerging Field Initiative (MRIC) (to MRF and TH) and from the French State in the frame of the ”Investments for the future” Programme IdEx Bordeaux, reference ANR-10-IDEX-03-02 (to MRF), by the National Institutes of Health (HL58984 and GM112455 to RB and NIH R01HL116571 to MX), Deutsche Forschungsgemeinschaft and Dr Werner Jackstädt-Stiftung (to MG), and from the American Heart Association (14POST18760003 to PKY). MW, MEW and RT are funded by the Medical Research Council (MR/M022706/1) and RT is the recipient of The Brian Ridge Scholarship. We are very grateful to Beatriz Alvarez (Universidad de la República, Montevideo, Uruguay) and Ivana Ivanovic-Burmazovic (Friedrich-Alexander University Erlangen-Nüremberg) for helpful discussions, suggestions and critical reading of the manuscript. We also thank Ody Sibon and Madina Baratashvili (University of Groningen) for D. melanogaster samples, and Andreas Gieβl and Johann Helmut Brandstätter (Friedrich-Alexander University Erlangen-Nüremberg) for the use of ChemiDoc detection system.
crosscheck: This document is CrossCheck deposited related_data: Supplementary Information copyright_licence: The Royal Society of Chemistry has an exclusive publication licence for this journal copyright_licence: This article is freely available. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence (CC BY 3.0) history: Received 13 December 2015; Accepted 9 February 2016; Accepted Manuscript published 9 February 2016
First published online 09 Feb 2016