Redox control of the DNA damage-inducible protein DinG helicase activity via its iron-sulfur cluster.
The Escherichia coli DNA damage-inducible protein DinG, a member of the superfamily 2 DNA helicases, has been implicated in the nucleotide excision repair and recombinational DNA repair pathways. Combining UV-visible absorption, EPR, and enzyme activity measurements, we demonstrate here that E. coli DinG contains a redox-active [4Fe-4S] cluster with a midpoint redox potential (E(m)) of -390 +/- 23 mV (pH 8.0) and that reduction of the [4Fe-4S] cluster reversibly switches off the DinG helicase activity. Unlike the [4Fe-4S] cluster in E. coli dihydroxyacid dehydratase, the DinG [4Fe-4S] cluster is stable, and the enzyme remains fully active after exposure to 100-fold excess of hydrogen peroxide, indicating that DinG could be functional under oxidative stress conditions. However, the DinG [4Fe-4S] cluster can be efficiently modified by nitric oxide (NO), forming the DinG-bound dinitrosyl iron complex with the concomitant inactivation of helicase activity in vitro and in vivo. Reassembly of the [4Fe-4S] cluster in NO-modified DinG restores helicase activity, indicating that the iron-sulfur cluster in DinG is the primary target of NO cytotoxicity. The results led us to propose that the iron-sulfur cluster in DinG may act as a sensor of intracellular redox potential to modulate its helicase activity and that modification of the iron-sulfur cluster in DinG and likely in other DNA repair enzymes by NO may contribute to NO-mediated genomic instability.
Ren
Binbin
B
Department of Biological Sciences, Louisiana State University, Baton Rouge, Louisiana 70803, USA.
Duan
Xuewu
X
Ding
Huangen
H
D004265:DNA Helicases / Q000737:chemistry* / Q000378:metabolism; D004260:DNA Repair / Q000502:physiology*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D042822:Genomic Instability / Q000502:physiology; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000737:chemistry* / Q000378:metabolism; D009569:Nitric Oxide / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000502:physiology; D013455:Sulfur / Q000737:chemistry* / Q000378:metabolism
D029968:Escherichia coli Proteins; C074432:dinG protein, E coli; D009569:Nitric Oxide; D013455:Sulfur; D006861:Hydrogen Peroxide; D007501:Iron; D004265:DNA Helicases