Comparison of the DNA-alkylating properties and mutagenic responses of a series of S-(2-haloethyl)-substituted cysteine and glutathione derivatives

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Abstract
The mutagenicity of 1,2-dibromoethane is highly dependent upon its conjugation of glutathione by the enzyme glutathione S-transferase. The conjugates thus formed can react with DNA and yield almost exclusively N7-guanyl adducts. We have synthesized the S-haloethyl conjugates of cysteine and glutathione, as well as selected methyl ester and N-acetyl derivatives, and compared them for ability to produce N7-guanyl adducts with calf thymus DNA. The cysteine compounds were found to be more reactive toward calf thymus DNA and yielded higher adduct levels than did the glutathione compounds. Adduct levels tended to be suppressed when there was a net charge on the compound and were not affected by substitution of bromine for chlorine, as expected for a mechanism known to involve an intermediate episulfonium ion. Sequence-selective alkylation of fragments of pBR322 DNA was investigated. The compounds produced qualitatively similar patterns of alkylation, with higher levels of alkylation at runs of guanines. The compounds were also tested for their ability to act as direct mutagens in Salmonella typhimurium TA98 and TA100. None of the compounds caused mutations in the TA98 frameshift mutagenesis assay. In the strain TA100, where mutation of a specific guanine by base-pair substitution produces reversion, all compounds were found to produce mutations, but the levels of mutagenicity did not correlate at all with the levels of DNA alkylation. The ratio of mutations to adducts varied at least 14-fold among the various N7-guanyl adducts examined. S-(2-Chloroethyl)glutathione was found to be the most potent mutagen, although it produced only intermediate levels of alkylation, indicating that the guanyl adduct formed in this case seems to be unusually mutagenic. The differences cannot be attributed to error-prone DNA repair mechanisms, since the activation of a umuC-containing plasmid (harbored in S. typhimurium TA 1535) was weak with all of the compounds examined and the response was in the opposite order of the mutation/adduct ratio. The results indicate that (1) N7-alkylguanine residues can be quite mutagenic and (2) small differences in the structures of a single such adduct can dramatically alter mutagenicity.