Abstract

Of the platinum complexes currently utilized in cancer chemotherapy oxaliplatin (OX) appears to be less mutagenic than cisplatin (CP) or carboplatin. The mechanism(s) for these differences is not clear, but their mutagenicity is likely to be determined in part by DNA polymerase(s) that catalyze translesion synthesis past Pt-DNA adducts and/or damage-recognition proteins that bind to the adducts and prevent translesion synthesis. We postulate that there are differences in conformation and/or conformational flexibility between CP and OX adducts that affect the efficiency and fidelity of translesion synthesis. We have recently obtained the NMR solution structure for the OX-GG adduct in the mutagenic AGG sequence context, and it is significantly different from all previous solution structures of the CP-GG adduct. Thus, we plan to solve the NMR solution structure of the CP adduct and conduct NMR relaxation studies on OX and CP adducts in the AGG sequence context (aim 1). Our in vitro studies show that pol eta, pol beta, and pol mu are the only DNA polymerases with a significant capacity to bypass CP and OX adducts. Our data show that pol eta expression decreases the mutagenicity of CP, and previous studies have shown that pol beta overexpression increases the mutagenicity of CP. Pol beta is an excellent model for studying translesion synthesis because of the wealth of available structural and kinetic information. HMG-domain proteins such as SRY bind to CP and OX adducts with different affinities and can block translesion synthesis. Thus, we plan to extract NMR structural and dynamic information for OX and CP adducts in complex with pol beta and the damage-recognition protein SRY (aim 2); utilize molecular dynamics to characterize the impact of CP and OX adducts on DNA flexibility and to characterize the interaction of these adducts with pol beta (aim 3); and characterize the pre-steady state kinetics of pol beta with templates containing OX and CP adducts (aim 4). These data will also be used to verify and refine the molecular dynamic simulations performed in aim 3. These experiments will help define the mechanism(s) of Pt drug-induced mutagenesis and will result in models of Pt-DNA-polymerase interactions that will be useful in clarifying the mechanisms of translesion synthesis past bulky DNA adducts.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA084480-06
Application #
7024962
Study Section
Cancer Etiology Study Section (CE)
Program Officer
Wolpert, Mary K
Project Start
1999-12-01
Project End
2009-02-28
Budget Start
2006-04-01
Budget End
2007-02-28
Support Year
6
Fiscal Year
2006
Total Cost
$288,183
Indirect Cost

  Institution

Name
University of North Carolina Chapel Hill
Department
Biochemistry
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599

  Publications

Ding, Feng; Lavender, Christopher A; Weeks, Kevin M et al. (2012) Three-dimensional RNA structure refinement by hydroxyl radical probing. Nat Methods 9:603-8
Bhattacharyya, Debadeep; Ramachandran, Srinivas; Sharma, Shantanu et al. (2011) Flanking bases influence the nature of DNA distortion by platinum 1,2-intrastrand (GG) cross-links. PLoS One 6:e23582
Hajdin, Christine E; Ding, Feng; Dokholyan, Nikolay V et al. (2010) On the significance of an RNA tertiary structure prediction. RNA 16:1340-9
Baskerville-Abraham, Irene M; Boysen, Gunnar; Troutman, J Mitchell et al. (2009) Development of an ultraperformance liquid chromatography/mass spectrometry method to quantify cisplatin 1,2 intrastrand guanine-guanine adducts. Chem Res Toxicol 22:905-12
Ramachandran, Srinivas; Temple, Brenda R; Chaney, Stephen G et al. (2009) Structural basis for the sequence-dependent effects of platinum-DNA adducts. Nucleic Acids Res 37:2434-48
Ding, Feng; Sharma, Shantanu; Chalasani, Poornima et al. (2008) Ab initio RNA folding by discrete molecular dynamics: from structure prediction to folding mechanisms. RNA 14:1164-73
Sharma, Shantanu; Ding, Feng; Dokholyan, Nikolay V (2008) iFoldRNA: three-dimensional RNA structure prediction and folding. Bioinformatics 24:1951-2
Chen, Yiwen; Ding, Feng; Nie, Huifen et al. (2008) Protein folding: then and now. Arch Biochem Biophys 469:4-19
Sharma, Shantanu; Gong, Peng; Temple, Brenda et al. (2007) Molecular dynamic simulations of cisplatin- and oxaliplatin-d(GG) intrastand cross-links reveal differences in their conformational dynamics. J Mol Biol 373:1123-40
Wu, Yibing; Bhattacharyya, Debadeep; King, Candice L et al. (2007) Solution structures of a DNA dodecamer duplex with and without a cisplatin 1,2-d(GG) intrastrand cross-link: comparison with the same DNA duplex containing an oxaliplatin 1,2-d(GG) intrastrand cross-link. Biochemistry 46:6477-87

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