Direct mapping of sites of modifications in DNA by tandem mass spectrometry

TOXI 14

Goutam Chowdhury, goutam.chowdhury@vanderbilt.edu, Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, 638 Robinson Research Building, 23rd and Pierce Avenues, Nashville, TN 37232 and F. Peter Guengerich, f.guengerich@vanderbilt.edu, Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University, 642 Robinson Research Building, 23rd and Pierce Aves, Nashville, TN 37232.
The tumorogenic potential of DNA lesions can depend on the nature and genomic position of the modifications generated by chemicals. Currently available methods for mapping DNA modifications require either cleavage or hydrolysis of the DNA, a problem with certain lesions. We have developed a “direct” LC-MS/MS method using collision-induced dissociation for detection and mapping sites of modification in DNA. The method does not require any prior DNA cleavage or hydrolysis and the use of radioactive or stable isotopes. Double-stranded DNA, representing a region of exon 5 from the p53 gene, was alkylated with benzo[a]pyrene diol epoxide and the adducts distribution was determined. Consistent with the literature the presence of 5-methylcytosine opposite the guanine at codon 157 increased the yield of alkylation. This method can be extended to other DNA modifications and mixtures of DNA fragments. (Supported by USPHS R01 ES10546 and P30 ES00264)
 

Young Investigator Session
8:15 AM-12:00 PM, Monday, August 20, 2007 BCEC -- 258C, Oral

Division of Chemical Toxicology

The 234th ACS National Meeting, Boston, MA, August 19-23, 2007