No abstract

We previously described a protein, isolated from human tissues and cells, that bound to a defined doublestranded oligonucleotide containing a single site-specifically placed 1,N'-ethenoadenine. It was further demonstrated that this protein was a glycosylase and released 1,N'6-ethenoadenine. We now rind that this enzyme also releases 3-methyladenine from methylated DNA and that 3-methyladenine-DNA glycosylase behaves in the same manner, binding to the ethenoadenine-containing oligonucleotide and cleaving both ethenoadenine and 3-methyladenine from DNA containing these adducts. The rate and extent ofglycosylase activities toward the two adducts are similar.Cyclic DNA adducts, such as 1,N6-ethenoadenine (EA), are produced by a variety of industrial and environmental chemicals. These include vinyl chloride (1, 2), urethane (3), vinyl carbamate (4), and other two-carbon compounds (5) metabolized in liver microsomes (5, 6). 3-Methyladenine (m3A) is a major product produced in DNA by methylating agents such as alkyl sulfates and nitrosoureas (7) and may also be formed in cells by nonenzymatic methylation by a normal constituent, S-adenosyl-L-methionine (8). The structures ofthe deoxyribonucleosides of EA and m3A are shown below. Repair of m3A, initiated by DNA glycosylases, occurs in organisms as diverse as bacteria and mammals and is well characterized (9). In addition, the cDNA encoding human m3A-DNA glycosylase has been cloned (10-12). This enzyme seems to have a fairly broad substrate range, including 3-alkyl-and 7-alkylpurines (13,14 MNU, N-methyl-N-nitrosourea; DMS, dimethyl sulfate; ANPG, alkyl-N-purine-DNA glycosylase; dsDNA, double-stranded DNA; ssDNA, single-stranded DNA. tTo whom reprint requests should be addressed.

N2,3-Ethenoguanine (N2,3-epsilon G) was recently identified in the liver of vinyl chloride-exposed rats. We have now synthesized the nucleoside and the 5'-diphosphate which was copolymerized with CDP. The deoxypolynucleotide complement, synthesized by AMV reverse transcriptase contained, in addition to dG, dC and dT. The total pyrimidine content was approximately equivalent to the N2,3-epsilon G content of the template. Incorporation of dC is neither lethal nor mutagenic, while dT incorporation represents a mutagenic event, occurring with approximately 20% frequency. N2,3-epsilon G X dT base pairs can have two hydrogen bonds with minimal helical distortion, as is also the case for N2,3-epsilon G X C base pairs. N2,3-epsilon G is the only derivative formed in vivo by the human carcinogen, vinyl chloride, that can be shown to have a high probability of causing transitions which could initiate malignant transformation.

Several different components contribute to the maintenance of 8-oxoG levels in human DNA, with the greatest contributor being the removal of 8-oxodGTP from the cellular nucleotide pool by hMTH1.