A specific partner for abasic damage in DNA
A specific partner for abasic damage in DNA
复制标题
作者：
Matray, TJ;Kool, ET
在大多数DNA复制模型中，沃森-克里克氢键驱动核苷酸掺入新的DNA链，并保持碱基与模板链的互补性。然而，对胸腺嘧啶和腺嘌呤的非极性类似物的研究表明，在没有氢键的情况下，复制仍然有效（1-4）。碱基对的复制也可能受到空间排阻的影响，因此插入的核苷酸需要具有正确的大小和形状，以使活性位点与模板碱基相匹配（5，6）。一个简单的空间排阻模型图不需要沃森-克里克氢键来解释复制的保真度，也不需要典型的嘌呤和嘧啶形状来酶促合成一个碱基对，如果每个碱基对都可以适合DNA双螺旋而没有空间应变（6）。在这里，我们测试这个想法，使用芘核苷三磷酸（dPTP），其中荧光的“基地”几乎是一样大的整个沃森克里克碱基对。我们表明，非氢键dPTP是有效的，特异性插入的DNA聚合酶相对的网站，缺乏DNA碱基。该过程的效率接近天然碱基对的效率，特异性为10（2）-10（4）倍。我们利用这些特性对DNA中的脱碱基损伤进行测序，这是体内DNA损伤的常见形式（7）。除了它们在识别这种遗传病变中的应用外，我们的研究结果表明，既不需要氢键也不需要嘌呤和嘧啶结构来形成具有高效率和选择性的碱基对。这些发现证实了空间互补性是DNA合成保真度的重要因素。
In most-models of DNA replication, Watson-Crick hydrogen bonding drives the incorporation of nucleotides into the new strand of DNA and maintains the complementarity of bases with the template strand. Studies with nonpolar analogues of thymine and adenine, however, have shown that replication is still efficient in the absence of hydrogen bonds(1-4). The replication of base pairs might also be influenced by steric exclusion, whereby inserted nucleotides need to be the correct size and shape to fit the active site against a template base(5,6). A simple steric-exclusion model map not require Watson-Crick hydrogen bonding to explain the fidelity of replication, nor should canonical purine and pyrimidine shapes be necessary for enzymatic synthesis of a base pair if each can fit into the DNA double helix without steric strain(6). Here we test this idea by using a pyrene nucleoside triphosphate (dPTP) in which the fluorescent 'base' is nearly as large as an entire Watson-Crick base pair. We show that the non-hydrogen-bonding dPTP is efficiently and specifically inserted by DNA polymerases opposite sites that lack DNA bases. The efficiency of this process approaches that of a natural base pair and the specificity is 10(2)-10(4)-fold. We use these properties to sequence abasic lesions in DNA, which are a common form of DNA damage in vivo(7). In addition to their application in identifying such genetic lesions, our results show that neither hydrogen bonds nor purine and pyrimidine structures are required to form a base pair with high efficiency and selectivity. These findings confirm that steric complementarity is an important factor in the fidelity of DNA synthesis.