Activity of the tetrapyrrole regulator CrtJ is controlled by oxidation of a redox active cysteine located in the DNA binding domain.
CrtJ from Rhodobacter capsulatus is a regulator of genes involved in the biosynthesis of haem, bacteriochlorophyll, carotenoids as well as structural proteins of the light harvesting-II complex. Fluorescence anisotropy-based DNA-binding analysis demonstrates that oxidized CrtJ exhibits ~20-fold increase in binding affinity over that of reduced CrtJ. Liquid chromatography electrospray tandem ionization mass spectrometric analysis using DAz-2, a sulfenic acid (-SOH)-specific probe, demonstrates that exposure of CrtJ to oxygen or to hydrogen peroxide leads to significant accumulation of a sulfenic acid derivative of Cys420 which is located in the helix-turn-helix (HTH) motif. In vivo labelling with 4-(3-azidopropyl)cyclohexane-1,3-dione (DAz-2) shows that Cys420 also forms a sulfenic acid modification in vivo when cells are exposed to oxygen. Moreover, a Cys420 to Ala mutation leads to a ~60-fold reduction of DNA binding activity while a Cys to Ser substitution at position 420 that mimics a cysteine sulfenic acid results in a ~4-fold increase in DNA binding activity. These results provide the first example where sulfenic acid oxidation of a cysteine in a HTH-motif leads to differential effects on gene expression.
Department of Molecular and Cellular Biochemistry, Indiana University, Bloomington, IN 47405, USA.
Zhuo
Z
Cheng
Jiang
J
Wu
Aaron
A
Setterdahl
Khalilah
K
Reddie
Kate
K
Carroll
Loubna A
LA
Hammad
Jonathan A
JA
Karty
Carl E
CE
Bauer
D001426:Bacterial Proteins / Q000378:metabolism*; D002853:Chromatography, Liquid; D003545:Cysteine / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019077:Helix-Turn-Helix Motifs; D006861:Hydrogen Peroxide / Q000378:metabolism; D008954:Models, Biological; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D011485:Protein Binding; D016354:Rhodobacter capsulatus / Q000201:enzymology*; D053719:Tandem Mass Spectrometry; D045725:Tetrapyrroles / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
D001426:Bacterial Proteins; C107183:CrtJ protein, Rhodobacter; D004268:DNA-Binding Proteins; D045725:Tetrapyrroles; D014157:Transcription Factors; D004247:DNA; D006861:Hydrogen Peroxide; D003545:Cysteine; D010100:Oxygen