This proposal aims to characterize the chemistry and biology associated with DNA-mediated charge transport (CT). Specifically, roles for DNA CT in how DNA is damaged and repaired under oxidative stress will be explored. Experiments are proposed to examine oxidative DNA damage at long range as well as DNA-mediated signaling to regulatory proteins. We will examine the distance range over which oxidative damage is funneled to a control sequence in mitochondria using rhodium photooxidants to generate damage at distant sites. Mitochondrial DNA mutations and their correlations with cancers will also be determined. We will further delineate the role of DNA CT in DNA damage detection by base excision repair (BER) enzymes that contain 4Fe-4S clusters. We will prepare a series of BER enzyme mutants to characterize the path for DNA/protein CT. CT will be probed using DNA electrochemistry, EPR, transient absorption spectroscopy and AFM. In vitro protein oxidations by guanine radicals in DNA will be generated by hole injection from tethered metallointercalators. Biological assays of repair in E. coli will test how effectively mutants of two BER enzymes, MutY and Endonuclease III (EndoIII), cooperate in detecting base lesions. Correlations will be drawn between this biological helper function, DNA/protein CT studies, and cancer predispositions in human MutY homologues. DNA-mediated CT for transcriptional regulation will also be examined in soxR, a transcriptional activator from E. coli containing an Fe-S cluster. We will test the initiation of transcription from a distance through long range oxidation of DNA-bound soxR using tethered photooxidants. Paralleling these CT studies with DNA-bound proteins, we will construct DNA assemblies containing tethered metallointercalators, cyclometallated complexes of Ir and Re(CO) diimine complexes, to characterize DNA-mediated reduction chemistry versus long range DNA- mediated oxidations.

Public Health Relevance

This proposal is focused on examining chemically how DNA is damaged and repaired when cells undergo oxidative stress. Damaged DNA, if unrepaired, leads to cancerous transformations. Understanding both the damage and repair process is critical to evaluating cancer predispositions.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM049216-18
Application #
8197925
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Anderson, Vernon
Project Start
1993-03-01
Project End
2013-11-30
Budget Start
2011-12-01
Budget End
2013-11-30
Support Year
18
Fiscal Year
2012
Total Cost
$366,270
Indirect Cost
$122,827
Name
California Institute of Technology
Department
Chemistry
Type
Schools of Engineering
DUNS #
009584210
City
Pasadena
State
CA
Country
United States
Zip Code
91125
Grodick, Michael A; Segal, Helen M; Zwang, Theodore J et al. (2014) DNA-mediated signaling by proteins with 4Fe-4S clusters is necessary for genomic integrity. J Am Chem Soc 136:6470-8
Arnold, Anna R; Barton, Jacqueline K (2013) DNA protection by the bacterial ferritin Dps via DNA charge transport. J Am Chem Soc 135:15726-9
Pheeney, Catrina G; Arnold, Anna R; Grodick, Michael A et al. (2013) Multiplexed electrochemistry of DNA-bound metalloproteins. J Am Chem Soc 135:11869-78
Sontz, Pamela A; Mui, Timothy P; Fuss, Jill O et al. (2012) DNA charge transport as a first step in coordinating the detection of lesions by repair proteins. Proc Natl Acad Sci U S A 109:1856-61
Barton, Jacqueline K; Olmon, Eric D; Sontz, Pamela A (2011) Metal Complexes for DNA-Mediated Charge Transport. Coord Chem Rev 255:619-634
Olmon, Eric D; Sontz, Pamela A; Blanco-Rodriguez, Ana Maria et al. (2011) Charge photoinjection in intercalated and covalently bound [Re(CO)3(dppz)(py)]+-DNA constructs monitored by time-resolved visible and infrared spectroscopy. J Am Chem Soc 133:13718-30
Genereux, Joseph C; Wuerth, Stephanie M; Barton, Jacqueline K (2011) Single-step charge transport through DNA over long distances. J Am Chem Soc 133:3863-8
Olmon, Eric D; Hill, Michael G; Barton, Jacqueline K (2011) Using metal complex reduced states to monitor the oxidation of DNA. Inorg Chem 50:12034-44
Romano, Christine A; Sontz, Pamela A; Barton, Jacqueline K (2011) Mutants of the base excision repair glycosylase, endonuclease III: DNA charge transport as a first step in lesion detection. Biochemistry 50:6133-45
Mui, Timothy P; Fuss, Jill O; Ishida, Justin P et al. (2011) ATP-stimulated, DNA-mediated redox signaling by XPD, a DNA repair and transcription helicase. J Am Chem Soc 133:16378-81

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