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DNA-Bound Redox Activity of DNA Repair Glycosylases Containing [4Fe-4S] Clusters

Boal, Amie K. and Yavin, Eylon and Lukianova, Olga A. and O'Shea, Valerie L. and David, Sheila S. and Barton, Jacqueline K. (2005) DNA-Bound Redox Activity of DNA Repair Glycosylases Containing [4Fe-4S] Clusters. Biochemistry, 44 (23). pp. 8397-8407. ISSN 0006-2960. https://resolver.caltech.edu/CaltechAUTHORS:20160229-103206847

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Abstract

MutY and endonuclease III, two DNA glycosylases from Escherichia coli, and AfUDG, a uracil DNA glycosylase from Archeoglobus fulgidus, are all base excision repair enzymes that contain the [4Fe-4S]^(2+) cofactor. Here we demonstrate that, when bound to DNA, these repair enzymes become redox-active; binding to DNA shifts the redox potential of the [4Fe-4S]^(3+/2+) couple to the range characteristic of high-potential iron proteins and activates the proteins toward oxidation. Electrochemistry on DNA-modified electrodes reveals potentials for Endo III and AfUDG of 58 and 95 mV versus NHE, respectively, comparable to 90 mV for MutY bound to DNA. In the absence of DNA modification of the electrode, no redox activity can be detected, and on electrodes modified with DNA containing an abasic site, the redox signals are dramatically attenuated; these observations show that the DNA base pair stack mediates electron transfer to the protein, and the potentials determined are for the DNA-bound protein. In EPR experiments at 10 K, redox activation upon DNA binding is also evident to yield the oxidized [4Fe-4S]^(3+) cluster and the partially degraded [3Fe-4S]1+ cluster. EPR signals at g = 2.02 and 1.99 for MutY and g = 2.03 and 2.01 for Endo III are seen upon oxidation of these proteins by Co(phen)_3^(3+) in the presence of DNA and are characteristic of [3Fe-4S]^(1+) clusters, while oxidation of AfUDG bound to DNA yields EPR signals at g = 2.13, 2.04, and 2.02, indicative of both [4Fe-4S]^(3+) and [3Fe-4S]^(1+) clusters. On the basis of this DNA-dependent redox activity, we propose a model for the rapid detection of DNA lesions using DNA-mediated electron transfer among these repair enzymes; redox activation upon DNA binding and charge transfer through well-matched DNA to an alternate bound repair protein can lead to the rapid redistribution of proteins onto genome sites in the vicinity of DNA lesions. This redox activation furthermore establishes a functional role for the ubiquitous [4Fe-4S] clusters in DNA repair enzymes that involves redox chemistry and provides a means to consider DNA-mediated signaling within the cell.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1021/bi047494nDOIArticle
http://pubs.acs.org/doi/abs/10.1021/bi047494nPublisherArticle
ORCID:
AuthorORCID
Boal, Amie K.0000-0002-1234-8472
David, Sheila S.0000-0001-5873-7935
Barton, Jacqueline K.0000-0001-9883-1600
Additional Information:© 2005 American Chemical Society. Received November 29, 2004; Revised Manuscript Received April 8, 2005. Publication Date (Web): May 18, 2005. Financial support for this work from the NIH (Grants GM49216 to J.K.B. and CA67985 to S.S.D.), the National Foundation for Cancer Research (J.K.B.), and the University of Utah Research Foundation (S.S.D.) is acknowledged. We thank A. Livingston for assistance in preparing enzymes, D. Ceres for technical assistance, and Prof. T. R. O’Connor for generous donation of endonuclease III. We also thank Dr. W. A. Franklin for providing the pET28aafung plasmid and Dr. M. P. Golinelli-Cohen for preparing the pMAL-c2X-mutY plasmid.
Funders:
Funding AgencyGrant Number
NIHGM49216
NIHCA67985
National Foundation for Cancer ResearchUNSPECIFIED
University of Utah Research FoundationUNSPECIFIED
Issue or Number:23
Record Number:CaltechAUTHORS:20160229-103206847
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20160229-103206847
Official Citation:DNA-Bound Redox Activity of DNA Repair Glycosylases Containing [4Fe-4S] Clusters Amie K. Boal, Eylon Yavin, Olga A. Lukianova, Valerie L. O'Shea, Sheila S. David, and Jacqueline K. Barton Biochemistry 2005 44 (23), 8397-8407 DOI: 10.1021/bi047494n
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:64846
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:29 Feb 2016 20:20
Last Modified:09 Mar 2020 13:18

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