Real-time imaging of the bacillithiol redox potential in the human pathogen Staphylococcus aureus using a genetically encoded bacilliredoxin-fused redox biosensor

Vu Van Loi, Manuela Harms, Marret Müller, Nguyen Thi Thu Huyen, Christopher J. Hamilton, Falko Hochgräfe, Jan Pane-Farre, Haike Antelmann

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Aims: Bacillithiol (BSH) is utilized as major thiol-redox buffer in the human pathogen Staphylococcus aureus. Under oxidative stress, BSH forms mixed disulfides with proteins, termed as S-bacillithiolation which can be reversed by bacilliredoxins (Brx). In eukaryotes, glutaredoxin-fused roGFP2 biosensors have been applied for dynamic live-imaging of the glutathione redox potential. Here, we have constructed a genetically encoded bacilliredoxin-fused redox biosensor (Brx-roGFP2) to monitor dynamic changes in the BSH redox potential in S. aureus.

Results: The Brx-roGFP2 biosensor showed a specific and rapid response to low levels bacillithiol disulphide (BSSB) in vitro which required the active-site Cys of Brx. Dynamic live-imaging in two methicillin-resistant S. aureus (MRSA) USA300 and COL strains revealed fast and dynamic responses of the Brx-roGFP2 biosensor under hypochlorite and H2O2 stress and constitutive oxidation of the probe in different BSH-deficient mutants. Furthermore, we found that the Brx-roGFP2 expression level and the dynamic range is higher in S. aureus COL compared to the USA300 strain. In phagocytosis assays with THP-1 macrophages, the biosensor was 87 % oxidized in S. aureus COL. However, no changes in the BSH redox potential were measured after treatment with different antibiotics classes indicating that antibiotics do not cause oxidative stress in S. aureus.

Conclusion and Innovation: This Brx-roGFP2 biosensor catalyzes specific equilibration between the BSH and roGFP2 redox couples and can be applied for dynamic live imaging of redox changes in S. aureus and other BSH-producing Firmicutes.
Original languageEnglish
Pages (from-to)835-848
Number of pages14
JournalAntioxidants & Redox Signaling
Issue number15
Early online date27 Jul 2016
Publication statusPublished - May 2017


  • Staphylococcus aureus
  • bacillithiol
  • bacilliredoxin
  • redox biosensor

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