8-oxo-2'-deoxyguanosine (OdG) is a prominent DNA lesion that can direct the incorporation of dCTP or dATP during replication. As the latter reaction can lead to mutation, the ratio of dCTP/dATP incorporation can significantly affect the mutagenic potential of OdG. Previous work with the A-family polymerase BF and seven analogues of OdG identified a major groove amino acid, Ile716, which likely influences the dCTP/dATP incorporation ratio opposite OdG. To further probe the importance of this amino acid, dCTP and dATP incorporations opposite the same seven analogues were tested with two BF mutants, I716M and I716A. Results from these studies support the presence of clashing interactions between Ile716 and the C8-oxygen and C2-amine during dCTP and dATP incorporations, respectively. Crystallographic analysis suggests that residue 716 alters the conformation of the template base prior to insertion into the active site, thereby affecting enzymatic efficiency. These results are also consistent with previous work with A-family polymerases, which indicate they have tight, rigid active sites that are sensitive to template perturbations.

中文翻译：

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Ile716对芽孢杆菌片段DNA聚合酶对8-Oxo-2'-脱氧鸟苷致突变性的重要性。

8-oxo-2'-脱氧鸟苷（OdG）是一个突出的DNA损伤，可指导复制过程中dCTP或dATP的掺入。由于后者反应会导致突变，因此dCTP / dATP掺入的比例会显着影响OdG的诱变潜力。以前使用A族聚合酶BF和OdG的七个类似物进行的研究确定了主要的沟纹氨基酸Ile716，这可能会影响与OdG相对的dCTP / dATP掺入率。为了进一步探讨该氨基酸的重要性，使用两个BF突变体I716M和I716A测试了与相同的七个类似物相对的dCTP和dATP掺入。这些研究的结果支持分别在dCTP和dATP掺入过程中Ile716与C8氧和C2-胺之间存在冲突的相互作用。晶体学分析表明，残基716在插入活性位点之前改变了模板碱基的构象，从而影响了酶促效率。这些结果也与先前使用A系列聚合酶的工作一致，这表明它们具有紧密，刚性的活性位点，这些活性位点对模板扰动敏感。