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Cysteine Reactivity Profiling to Unveil Redox Regulation in Phytopathogens

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Plant Proteases and Plant Cell Death

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2447))

Abstract

Reactivity-based chemical proteomics is a powerful technology based on the use of tagged chemicals that covalently react with surface-exposed residues on proteins in native proteomes. Reactivity profiling involves the purification, identification, and quantification of labeled peptides by LC-MS/MS. Here, we have detailed a protocol for reactivity profiling of Cys residues using iodoacetamide probes, displaying >1000 reactive Cys residues in the proteome of phytopathogen Pseudomonas syringae pv. tomato DC3000 (PtoDC3000). Comparative reactivity profiling of PtoDC3000 treated with or without hydrogen peroxide (H2O2) identified ~200 H2O2-sensitive Cys residues in antioxidant enzymes, metabolic enzymes, and transcription regulators. Interestingly, half of these H2O2-sensitive Cys residues are more reactive in response to H2O2 and several proteins have multiple Cys residues with opposite reactivities in response to H2O2 exposure.

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Acknowledgments

This research was supported by the John Fell Fund.

Author information

Authors and Affiliations

  1. The Plant Chemetics Laboratory, Department of Plant Sciences, University of Oxford, Oxford, UK

    Kyoko Morimoto & Renier A. L. van der Hoorn

  2. Advanced Proteomics Centre, Department of Biochemistry, University of Oxford, Oxford, UK

    Monika Stegmann

  3. ZMB Chemical Biology, Faculty of Biology, University of Duisburg-Essen, Essen, Germany

    Farnusch Kaschani

  4. Department of Chemistry, Chemistry Research Laboratory, Oxford, UK

    Shabaz Mohammed

  5. Department of Biochemistry, New Biochemistry Building, Oxford, UK

    Shabaz Mohammed

  6. The Rosalind Franklin Institute, Oxfordshire, UK

    Shabaz Mohammed

Authors
  1. Kyoko Morimoto
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  2. Monika Stegmann
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  3. Farnusch Kaschani
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  4. Shabaz Mohammed
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  5. Renier A. L. van der Hoorn
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Corresponding author

Correspondence to Renier A. L. van der Hoorn .

Editor information

Editors and Affiliations

  1. Chemistry and Chemical Technology, University of Ljubljana, Ljubljana, Slovenia

    Marina Klemenčič

  2. VIB-UGent Cntr for Plant Systems Biology, Ghent University, Ghent, Belgium

    Simon Stael

  3. ZEA-3 Analytik, Forschungszentrum Juelich, Juelich, Germany

    Pitter F. Huesgen

1 Electronic Supplementary Material

Table S1

Oligonucleotides used for Darwin Assembly and Barcoding Libraries (XLSX 472 kb)

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© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Morimoto, K., Stegmann, M., Kaschani, F., Mohammed, S., van der Hoorn, R.A.L. (2022). Cysteine Reactivity Profiling to Unveil Redox Regulation in Phytopathogens. In: Klemenčič, M., Stael, S., Huesgen, P.F. (eds) Plant Proteases and Plant Cell Death. Methods in Molecular Biology, vol 2447. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2079-3_9

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  • DOI: https://doi.org/10.1007/978-1-0716-2079-3_9

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2078-6

  • Online ISBN: 978-1-0716-2079-3

  • eBook Packages: Springer Protocols

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