Melchior W B, Marques M M, Beland F A
Division of Biochemical Toxicology, National Center for Toxicological Research, Jefferson, AR 72079.
Carcinogenesis. 1994 May;15(5):889-99. doi: 10.1093/carcin/15.5.889.
A 276 bp region from the tetracycline resistance gene of the plasmid pBR322 was modified with 2-acetylaminofluorene (AAF), 2-aminofluorene (AF), 4-aminobiphenyl (ABP), N'-acetylbenzidine or 1-aminopyrene (AP) in order to determine the effect of adduct structure upon mutation induction. Each modification reaction gave one major adduct and these adducts had chromatographic properties, as determined by 32P-postlabeling, identical to those in which substitution had occurred at C8 of deoxyguanosine through the amine or amide nitrogen. The types and distribution of mutations were then characterized following introduction of the modified plasmids into SOS-induced Escherichia coli using Hanahan et al.'s procedure (Methods Enzymol., 204, 63-113, 1991). With AAF-modified plasmid, 60% of the mutations were deletions or additions, and these were detected primarily at NarI sites or in repetitive G sequences. Modification with AF gave -G deletions, primarily in runs of Gs, and base substitution mutations, which were mainly G to T transversions. Substitution with ABP or N'-acetylbenzidine resulted in G to T and G to C transversions, the latter being a mutation not detected with AF; in addition, -G deletions were detected at only very low frequency. AP modification gave both -G frameshift and base substitution mutations, of which G to T transversions predominated. A comparison of the mutation frequencies per adduct indicated that the mutagenic efficiencies of the adducts decreased in the order AP > AF > AAF approximately ABP approximately N'-acetylbenzidine. AAF- and ABP-modified pBR322 were also introduced with a CaCl2 method. The mutation frequency per adduct increased with this transformation procedure, and this appeared to be a reflection of a greater percentage of frameshift mutations. These data indicate that a series of structurally related aromatic amines will induce both base substitution and frameshift mutations when incorporated into pBR322, but that frameshift mutations occur almost exclusively with the planar derivatives. Furthermore, the ability to induce frameshift mutations increases the mutagenic efficiency of an adduct.
为了确定加合物结构对突变诱导的影响，用2-乙酰氨基芴（AAF）、2-氨基芴（AF）、4-氨基联苯（ABP）、N'-乙酰联苯胺或1-氨基芘（AP）对质粒pBR322的四环素抗性基因的一段276 bp区域进行了修饰。每个修饰反应都产生一种主要加合物，通过32P后标记法测定，这些加合物具有与通过胺或酰胺氮在脱氧鸟苷的C8处发生取代的加合物相同的色谱性质。然后按照哈纳汉等人的方法（《酶学方法》，204，63 - 113，1991），将修饰后的质粒导入SOS诱导的大肠杆菌后，对突变的类型和分布进行了表征。用AAF修饰的质粒，60%的突变是缺失或插入，主要在NarI位点或重复的G序列中检测到。用AF修饰产生了-G缺失，主要在G的连续序列中，以及碱基替代突变，主要是G到T的颠换。用ABP或N'-乙酰联苯胺替代导致了G到T和G到C的颠换，后者是用AF未检测到的突变；此外，-G缺失仅在非常低的频率下检测到。AP修饰产生了-G移码和碱基替代突变，其中G到T的颠换占主导。对每个加合物的突变频率进行比较表明，加合物的诱变效率按AP > AF > AAF ≈ ABP ≈ N'-乙酰联苯胺的顺序降低。AAF和ABP修饰的pBR322也用氯化钙法导入。每个加合物的突变频率随着这种转化方法而增加，这似乎反映了移码突变的比例更高。这些数据表明，一系列结构相关的芳香胺在掺入pBR322时会诱导碱基替代和移码突变，但移码突变几乎只发生在平面衍生物中。此外，诱导移码突变的能力提高了加合物的诱变效率。
