Sequence-specific DNA interstrand cross-linking by an aziridinomitosene in the absence of exogenous reductant.
Sequence-specific DNA interstrand cross-linking by an aziridinomitosene in the absence of exogenous reductant.
复制标题
在没有外源还原剂的情况下，通过氮丙啶核丝烯进行序列特异性 DNA 链间交联。
DOI：
10.1021/bi050426w
复制
发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Vedejs,Edwin
中科院分区：
文献类型：
--
作者：
Rink,StaciaM;Warner,DonL;Klapars,Artis;Vedejs,Edwin
氮丙啶基核糖烯衍生物 (1S,2S)-6-去甲基（甲基氮丙啶基）核糖烯 (4) 在 pH 7.4 下，在没有还原剂的情况下可烷基化质粒 DNA [Vedejs, E., Naidu, B. N., Klapars, A., Warner, D. L., Li, V. -s., Na, Y., and Kohn, H. (2003) J.是。化学。 Soc.125, 15796−15806]，母体丝裂霉素中未发现的活性。我们试图使用非还原反应条件评估氮丙啶基线粒体 DNA 链间交联活性的存在。用 4 处理的放射性标记 DNA 通过变性聚丙烯酰胺凝胶电泳 (DPAGE) 进行分析，该技术可以轻松地将移动性较差的交联双链 DNA 与移动性较高的单链 DNA 产物分开。 Nonreduced4 在含有 5'-d(CG) 位点的双链 DNA 中产生链间交联 (ICL)，并且 ICL 的产量与相似条件下从还原 MC 获得的产量相当。具有中心四核苷酸 5'-d(ACGT) 的双链 DNA 提供来自非还原 4 和还原 MC 的最大 ICL 产量。用反向序列 5'-d(GC) 替换 5'-d(CG) 完全消除了 4 的链间交联，揭示了 5'-d(CG) 是 ICL 形成的特定位点。将 5'-d(CG) 处的 dG 替换为 2'-脱氧肌苷 (dI)（缺乏 dG 中存在的环外 C2 氨基），也阻止了 4 形成 DNA ICL，揭示了 dG C2 氨基在 4 和双链 DNA 之间的链间交联反应中的重要作用。该报告直接证明了非还原氮丙啶基核糖烯中存在双功能烷基化活性，并清楚地表明未还原 4 烷基化 ds DNA 小沟中的残基，与还原 MC 显示的相同 5'-d(CG) 序列特异性交联。
The aziridinomitosene derivative (1S,2S)-6-desmethyl(methylaziridino)mitosene (4) was shown to alkylate plasmid DNA at pH 7.4in the absence of a reducing agent[Vedejs, E., Naidu, B. N., Klapars, A., Warner, D. L., Li, V. -s., Na, Y., and Kohn, H. (2003)J. Am. Chem. Soc.125, 15796−15806], an activity not found in the parent mitomycins. We sought to evaluate aziridinomitosene4for the presence of DNA interstrand cross-linking activity using nonreductive reaction conditions. Radiolabeled DNA treated with4was analyzed by denaturing polyacrylamide gel electrophoresis (DPAGE), a technique that readily separates the less mobile cross-linked ds DNA from the more mobile ss DNA products. Nonreduced4produced an interstrand cross-link (ICL) in duplex DNA containing 5‘-d(CG) sites, and the yield of ICL was comparable to that obtained from reduced MC under similar conditions. A ds DNA having the central tetranucleotide 5‘-d(ACGT) provided the greatest ICL yield from both nonreduced4and reduced MC. Substitution of 5‘-d(CG) with the inverted sequence 5‘-d(GC) completely abolished interstrand cross-linking by4, revealing 5‘-d(CG) as its specific site of ICL formation. Replacement of dG at 5‘-d(CG) with 2‘-deoxyinosine (dI), which lacks the exocyclic C2amino group present in dG, also prevented DNA ICL formation by4, revealing an essential role for the dG C2amino group in the interstrand cross-linking reaction between4and duplex DNA. This report directly demonstrates the presence of bifunctional alkylating activity in a nonreduced aziridinomitosene and clearly shows that unreduced4alkylates residues in the minor groove of ds DNA, cross-linking with the same 5‘-d(CG) sequence specificity displayed by reduced MC.