Mutagenic potential of guanine N2 adducts of butadiene mono- and diolepoxide
Mutagenic potential of guanine N2 adducts of butadiene mono- and diolepoxide
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DOI：
10.1021/tx9901332
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发表时间：
2000-01-01
影响因子：
4.1
通讯作者：
Lloyd, RS
中科院分区：
文献类型：
--
作者：
Carmical, JR;Zhang, MZ;Lloyd, RS
为了探索立体异构丁二烯代谢物的鸟嘌呤 N-2 加合物在丁二烯诱导的诱变中的作用，制备了含有 (R)- 和 (S)- 单环氧化物以及 (RP)- 和 (S,S)- 二烯环氧化物的加合物的 II-mer 脱氧寡核苷酸。这些加合的寡核苷酸被用于体内和体外实验，旨在检查每种寡核苷酸的诱变效力及其通过大肠杆菌聚合酶的复制。将四个加合的脱氧寡核苷酸中的每一个连接到单链M13mp7L2载体中并转染到大肠杆菌中。筛选所得噬菌斑是否存在 N-ras 12 密码子第 2 位的错误掺入。尽管诱变频率较低，但各种立体异构体的不同相对诱变性是可辨别的。此外，通过引物延伸测定确定了每种加合物对三种主要大肠杆菌聚合酶的生物效应。加合的 11 聚体连接到 60 聚体线性 DNA 分子中，为引物延伸提供足够长的模板。通过引物延伸测定，所有四种鸟嘌呤加合物被确定对三种聚合酶中的每一种都具有阻断作用。
To explore the role of guanine N-2 adducts of st;stereoisomeric bu;diene metabolites in butadiene-induced mutagenesis, Il-mer deoxyoligonucleotides were prepared containing adducts of (R)- and (S)-monoepoxide and (RP)- and (S,S)-diolepoxide. These adducted oligonucleotides were utilized in both in vivo and in vitro experiments designed to examine the mutagenic potency of each and their replication by Escherichia coli polymerases. Each of the four adducted deoxyoligonucleotides was ligated into a single-stranded M13mp7L2 vector and transfected into E. coli. The-resulting plaques were screened for misincorporation at position 2 of the N-ras 12 codon. Although the mutagenic frequencies were low, different relative mutagenicities of the various stereoisomers were discernible. In addition, the biological effects of each adduct on the three major E. coli polymerases were determined via primer extension assays. The adducted 11-mers were ligated into a 60-mer linear DNA molecule to provide a sufficiently long template for primer elongation. All four guanine adducts were determined to be blocking to each of the three polymerases via primer extension assays.