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The Oxidation State of [4Fe4S] Clusters Modulates the DNA-Binding Affinity of DNA Repair Proteins

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journal contribution
posted on 17.08.2017, 00:00 by Edmund C. M. Tse, Theodore J. Zwang, Jacqueline K. Barton
A central question important to understanding DNA repair is how certain proteins are able to search for, detect, and fix DNA damage on a biologically relevant time scale. A feature of many base excision repair proteins is that they contain [4Fe4S] clusters that may aid their search for lesions. In this paper, we establish the importance of the oxidation state of the redox-active [4Fe4S] cluster in the DNA damage detection process. We utilize DNA-modified electrodes to generate repair proteins with [4Fe4S] clusters in the 2+ and 3+ states by bulk electrolysis under an O2-free atmosphere. Anaerobic microscale thermophoresis results indicate that proteins carrying [4Fe4S]3+ clusters bind to DNA 550 times more tightly than those with [4Fe4S]2+ clusters. The measured increase in DNA-binding affinity matches the calculated affinity change associated with the redox potential shift observed for [4Fe4S] cluster proteins upon binding to DNA. We further devise an electrostatic model that shows this change in DNA-binding affinity of these proteins can be fully explained by the differences in electrostatic interactions between DNA and the [4Fe4S] cluster in the reduced versus oxidized state. We then utilize atomic force microscopy (AFM) to demonstrate that the redox state of the [4Fe4S] clusters regulates the ability of two DNA repair proteins, Endonuclease III and DinG, to bind preferentially to DNA duplexes containing a single site of DNA damage (here a base mismatch) which inhibits DNA charge transport. Together, these results show that the reduction and oxidation of [4Fe4S] clusters through DNA-mediated charge transport facilitates long-range signaling between [4Fe4S] repair proteins. The redox-modulated change in DNA-binding affinity regulates the ability of [4Fe4S] repair proteins to collaborate in the lesion detection process.