DNA−Protein Cross-Linking by 1,2,3,4-Diepoxybutane

Abstract

Abstract1,2,3,4-Diepoxybutane (DEB) is a strongly genotoxic diepoxide hypothesized to be the ultimate carcinogenic metabolite of the common industrial chemical and environmental carcinogen 1,3-butadiene. DEB is a bis-electrophile capable of cross-linking cellular biomolecules to form DNA-DNA and DNA-protein cross-links (DPCs), which are thought to play a central role in its biological activity. Previous studies with recombinant proteins have shown that the biological outcomes of DEB-induced DPCs are strongly influenced by protein identities. The present work combines affinity capture methodology with mass spectrometry-based proteomics and immunological detection to identify the proteins which form DPCs in nuclear extracts from human cervical carcinoma (HeLa) cells. We identified 39 human proteins that form covalent DPCs in the presence of DEB. DNA-protein cross-linking efficiency following treatment with 25 mM DEB was 2-12%, depending on protein identity. HPLC-ESI + -MS/MS analysis of the 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.

Bifunctional Electrophiles Cross-Link Thioredoxins with Redox Relay Partners in Cells

“…32,33,40-43 These molecules have a number of molecular targets and, in some instances, mediate inter-strand cross-linking within DNA and cross-linking between DNA and proteins. 44-48 In addition, our studies suggest that they may bring about some of their toxic effects by cross-linking proteins to one another, including those proteins in the Trx relay network. In the latter scenario, these electrophiles can directly form adducts with the redox active Cys residues in Trx relay proteins, thereby disrupting this key oxidant defense pathway and potentially promoting redox imbalance.…”

Replication past the Butadiene Diepoxide-Derived DNA Adduct S-[4-(N6-Deoxyadenosinyl)-2,3-dihydroxybutyl]glutathione by DNA Polymerases

“…11,12 Among the three epoxide metabolites of BD, DEB is the most mutagenic (50- to 100-fold > EB and 100- to 200-fold > 3,4-epoxy-1,2-butanediol), 13,14 presumably because of its bis electrophilic properties and enhanced reactivity with DNA. 14,15 Many DNA adducts derived from DEB have been reported, including mono-adducts and DNA-DNA intra- and inter-crosslinks adducts (≥ 25 adducts, with the number being even higher when stereochemistry is considered), 6–21 and some of these adducts have been found in vivo . 16,17 …”

Bypass of DNA-Protein Cross-links Conjugated to the 7-Deazaguanine Position of DNA by Translesion Synthesis Polymerases

“…Covalent entrapment of cellular proteins on genomic DNA is a common process that occurs upon exposure to a variety of endogenous and exogenous bis-electrophiles, heavy metals, and free radicals (1)(2)(3)(4)(5)(6)(7). Specifically, common chemotherapeutics such as nitrogen mustards, platinum compounds, and alkylnitrosoureas (3,4,8); environmental carcinogens such as formaldehyde and 1,3-butadiene (5, 9, 10); toxic metals (11)(12)(13); nitric oxide (14); free radicals (15,16); UV light (17); and ionizing radiation (6) mediate the formation of covalent DNAprotein cross-links (DPCs).…”

“…The most common site on DNA that participates in DNAprotein cross-linking mediated by bis-electrophiles is the N7 atom of guanine (3)(4)(5)10). However, to our knowledge, no reports are available in the literature in regard to the replication bypass of N7-guanine adducts, probably because of their labile nature.…”