Translesion synthesis by Sulfolobus solfataricus DNA polymerase Dpo4 across polycyclic aromatic hydrocarbon diol epoxide adducts of deoxyadenosine

TOXI 30

Goutam Chowdhury, goutam.chowdhury@vanderbilt.edu1, Hong Zang, hong.zang@vanderbilt.edu1, Karen C. Angel1, Thomas M Harris, thomas.h.harris@vanderbilt.edu2, and F. Peter Guengerich1. (1) Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, 638 Robinson Research Building, 23rd and Pierce Avenues, Nashville, TN 37232, (2) Department of Chemistry, Center in Molecular Toxicology and the Vanderbilt-Ingram Cancer Center, Vanderbilt University, 7332 Stevenson Center, Nashville, TN 37235
Polycyclic aromatic hydrocarbons (PAHs) are potent environmental carcinogens known over hundred years. PAHs are metabolically activated to reactive diol epoxides which react with DNA to form various covalent adducts. The mechanisms by which DNA adducts of PAH-derivatives cause mutations have been of immense interest. Three different N6-adenyl PAH-diol epoxide oligonucleotide derivatives (dA-ATBA-1S, dA-ATBA-11S and dA-STBP-10S) were studied with the archebacterial translesion DNA polymerase Sulfolobus solfataricus Dpo4. Steady-state kinetic analysis indicated insertion of all four dNTPs opposite all three N6-adenyl PAH adducts, with only slightly varying misincorporation efficiencies. Full-length extension across the N6-adenyl PAH derivatives proceeded to apparent completion at 45 °C in the presence of added dimethylsulfoxide. Analysis of the products by LCMS/MS indicated the presence of mixtures of products corresponding to both error-free synthesis and mixtures of polymerization/realignment steps. The results demonstrate the complexity of polymerization opposite these bulky N6-adenyl PAH adducts, even with a single polymerase.

 

General Papers: Young Investigator Session
8:00 AM-12:00 PM, Tuesday, 12 September 2006 Moscone Center -- Room 308, Oral

Sci-Mix
8:00 PM-10:00 PM, Monday, 11 September 2006 Moscone Center -- Hall D, Sci-Mix

Division of Chemical Toxicology

The 232nd ACS National Meeting, San Francisco, CA, September 10-14, 2006