Nitric Oxide pp 183-203 | Cite as

Identification of NO-Sensitive Cysteine Residues Using Cysteine Mutants of Recombinant Proteins

  • Azam Shekariesfahlan
  • Christian Lindermayr
Part of the Methods in Molecular Biology book series (MIMB, volume 1747)


Nitric oxide (NO) is a free radical gas regulating a wide range of biological processes in plants. Proteins are the main reaction target of NO inside the cells. The relevance of S-nitrosation as one of the NO-mediated protein posttranslational modifications has been studied in detail. S-nitrosylation causes alterations of the activity/function, sub-cellular localization or interaction partners of proteins. Up to present, a large number of S-nitrosation candidates have been detected in plants. Recombinant proteins are widely used to show or confirm the protein posttranslational modifications. Here, using recombinant proteins subjected to biotin switch assay, the S-nitrosation of some nuclear candidates of Arabidopsis is verified. Proteins usually contain several cysteine residues which each might involve in structure of protein active sites. So, an important question is: which cysteine residue is the target of S-nitrosation and does it belong to an active site? Here, using the approach of substitution of cysteines by serines on recombinant proteins, the NO-sensitive cysteine residue of an Arabidopsis nuclear protein is identified. The next step could be to investigate the effect of S-nitrosation on protein activity/function and further to test the role of target cysteines and S-nitrosation of them in protein activity/function.

Key words

Nitric oxide S-nitrosation Biotin switch assay Arabidopsis nuclear proteins Recombinant proteins Cysteine mutation 


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Copyright information

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

Authors and Affiliations

  • Azam Shekariesfahlan
    • 1
  • Christian Lindermayr
    • 2
  1. 1.Iranian Research Institute of Plant ProtectionAgricultural Research, Education and Extension Organization (AREEO)TehranIran
  2. 2.Helmholtz Zentrum München–German Research GmbH, Center for Environment HealthInstitute of Biochemical Plant Pathology, Ingolstädter Landstraße 1Munich-NeuherbergGermany

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