DNA Oxidation by Charge Transport in Mitochondria
- Creators
- Merino, Edward J.
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Barton, Jacqueline K.
Abstract
Sites of oxidative DNA damage in functioning mitochondria have been identified using a rhodium photooxidant as a probe. Here we show that a primer extension reaction can be used to monitor oxidative DNA damage directly in functioning mitochondria after photoreaction with a rhodium intercalator that penetrates the intact mitochondrial membrane. The complex [Rh(phi)_2bpy]Cl_3 (phi = 9,10-phenanthrenequinonediimine) binds to DNA within the mitochondria and, upon irradiation, initiates DNA oxidation reactions. Significantly, piperidine treatment of the mitochondria leads to protein-dependent primer extension stops spaced every ∼20 base pairs. Hence, within the mitochondria, the DNA is well covered and packaged by proteins. Photolysis of the mitochondria containing [Rh(phi)_2bpy]^(3+) leads to oxidative DNA damage at positions 260 and 298; both are mutational hot spots associated with cancers. The latter position is the 5'-nucleotide of conserved sequence block II and is critical to replication of the mitochondrial DNA. The oxidative damage is found to be DNA-mediated, utilizing a charge transport mechanism, as the Rh binding sites are spatially separated from the oxidation-prone regions. This long-range DNA-mediated oxidation occurs despite protein association. Indeed, the oxidation of the mitochondrial DNA leads not only to specific oxidative lesions, but also to a corresponding change in the protein-induced stops in the primer extension. Mitochondrial DNA damage promotes specific changes in protein−DNA contacts and is thus sensed by the mitochondrial protein machinery.
Additional Information
© 2008 American Chemical Society. Received August 30, 2007; Revised Manuscript Received November 20, 2007. Publication Date (Web): January 12, 2008. We are grateful to the NIH (Grant GM49216) for their financial support of this research including a minority postdoctoral fellowship to E.J.M.Attached Files
Supplemental Material - bi701775s_si_002.pdf
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Additional details
- Eprint ID
- 64852
- DOI
- 10.1021/bi701775s
- Resolver ID
- CaltechAUTHORS:20160229-120833212
- NIH
- GM49216
- NIH Postdoctoral Fellowship
- Created
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2016-02-29Created from EPrint's datestamp field
- Updated
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2021-11-10Created from EPrint's last_modified field