Reactive oxygen species induced by ionizing radiation and metabolic pathways generate 7,8-dihydro-8-oxoguanine (oxoG) and 7,8-dihydro-8-oxoadenine (oxoA) as two major forms of oxidative damage. The mutagenicity of oxoG, which promotes G to T transversions, is attributed to the lesion's conformational flexibility that enables Hoogsteen base pairing with dATP in the confines of DNA polymerases. The mutagenesis mechanism of oxoA, which preferentially causes A to C transversions, remains poorly characterized. While structures for oxoA bypass by human DNA polymerases are available, that of prokaryotic DNA polymerases have not been reported. Herein, we report kinetic and structural characterizations of Sulfolobus solfataricus Dpo4 incorporating a nucleotide opposite oxoA. Our kinetic studies show oxoA at the templating position reduces the replication fidelity by ∼560-fold. The catalytic efficiency of the oxoA:dGTP insertion is ∼300-fold greater than that of the dA:dGTP insertion, highlighting the promutagenic nature of oxoA. The relative efficiency of the oxoA:dGTP misincorporation is ∼5-fold greater than that of the oxoG:dATP misincorporation, suggesting the mutagenicity of oxoA is comparable to that of oxoG. In the Dpo4 replicating base pair site, oxoA in the anti-conformation forms a Watson-Crick base pair with an incoming dTTP, while oxoA in the syn-conformation assumes Hoogsteen base pairing with an incoming dGTP, displaying the dual coding potential of the lesion. Within the Dpo4 active site, the oxoA:dGTP base pair adopts a Watson-Crick-like geometry, indicating Dpo4 influences the oxoA:dGTP base pair conformation. Overall, the results reported here provide insights into the miscoding properties of the major oxidative adenine lesion during translesion synthesis.

中文翻译：

---

跨病变合成DNA聚合酶Dpo4促突变的7,8-二氢-8-氧腺嘌呤旁路。

电离辐射和代谢途径诱导的活性氧产生7,8-dihydro-8-oxoguanine（oxoG）和7,8-dihydro-8-oxoadenine（oxoA）是氧化损伤的两种主要形式。oxoG的诱变性，促进了G向T的转化，归因于病变的构象柔韧性，使DNA聚合酶范围内Hoogsteen碱基与dATP配对。oxoA的诱变机制，其优先引起A到C的转变，仍然缺乏明确的特征。虽然可以利用人DNA聚合酶绕过oxoA的结构，但尚未报道原核DNA聚合酶的结构。在这里，我们报告了Sulfolobus solfataricus Dpo4结合一个与oxoA相反的核苷酸的动力学和结构特征。我们的动力学研究表明，在模板位置上的oxoA使复制保真度降低了约560倍。oxoA：dGTP插入的催化效率比dA：dGTP插入的催化效率高约300倍，这突出了oxoA的致突变性。oxoA：dGTP错掺的相对效率比oxoG：dATP错掺的相对效率高约5倍，这表明oxoA的诱变性与oxoG相当。在Dpo4复制碱基对位点中，反构象中的oxoA与输入的dTTP形成沃森-克里克碱基对，而同构象中的oxoA则假定Hoogsteen碱基与输入的dGTP配对，显示了病变的双重编码潜力。在Dpo4活性位点内，oxoA：dGTP碱基对采用类似Watson-Crick的几何形状，表明Dpo4影响oxoA：dGTP碱基对构象。总体而言，此处报道的结果提供了跨病变合成过程中主要氧化腺嘌呤病变的误编码特性的见解。