1,2,3,4-diepoxybutane  (DEB)  is a key carcinogenic metabolite of 1,3-butadiene (BD), an important industrial and environmental chemical present in urban air and in cigarette smoke. DEB is a bis-electrophile capable of cross-linking cellular biomolecules to form DNA-DNA and DNA-protein cross-links (DPCs). In the present work, a mass spectrometry-based proteomics and immunological detection was employed to characterize DEB-mediated DNA-protein cross-linking in human fibrosarcoma (HT1080) cells. More than 60 proteins that were covalently cross-linked to chromosomal DNA, with a variety of biological functions were identified with high confidence. HPLC-ESI+-MS/MS analysis of total proteolytic digests of cross-linked proteins revealed the presence of 1-(S-cysteinyl)-4-(guan-7-yl)-2,3-butanediol conjugates, suggesting that DEB forms DPCs between cysteine thiols within proteins and the N-7 guanine positions within DNA. Synthetic protein monoepoxides introduced into HT1080 cells by electroporation caused mutations and cell death, suggesting that super-bulky DPC lesions contribute to the cytotoxic and mutagenic effects of DEB.

Characterization of DNA-protein cross-linking of 1,2,3,4-diepoxybutane by  mass spectrometry-based proteomics

244th National Meeting (2012)

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