Utilizing redox-sensitive GFP fusions to detect in vivo redox changes in a genetically engineered prokaryote

Wilhad Hans Reuter, Thorsten Masuch, Na Ke, Marine Lenon, Meytal Radzinski, Vu Van Loi, Guoping Ren, Paul Riggs, Haike Antelmann, Dana Reichmann, Lars I. Leichert, Mehmet Berkmen*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

Understanding the in vivo redox biology of cells is a complex albeit important biological problem. Studying redox processes within living cells without physical disruption or chemical modifications is essential in determining the native redox states of cells. In this study, the previously characterized reduction-oxidation sensitive green fluorescent protein (roGFP2) was used to elucidate the redox changes of the genetically engineered Escherichia coli strain, SHuffle. SHuffle cells were demonstrated to be under constitutive oxidative stress and responding transcriptionally in an OxyR-dependent manner. Using roGFP2 fused to either glutathione (GSH)- or hydrogen peroxide (H2O2)- sensitive proteins (glutaredoxin 1 or Orp1), the cytosolic redox state of both wild type and SHuffle cells based on GSH/GSSG and H2O2 pools was measured. These probes open the path to in vivo studies of redox changes and genetic selections in prokaryotic hosts.

Original languageAmerican English
Article number101280
JournalRedox Biology
Volume26
DOIs
StatePublished - Sep 2019

Bibliographical note

Publisher Copyright:
© 2019 The Authors

Keywords

  • Disulfide bond formation
  • Redox
  • SHuffle
  • roGFP

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