Role of hoogsteen edge hydrogen bonding at template purines in nucleotide incorporation by human DNA polymerase iota.
Role of hoogsteen edge hydrogen bonding at template purines in nucleotide incorporation by human DNA polymerase iota.
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模板嘌呤上的 Hoogsteen 边缘氢键在人类 DNA 聚合酶 iota 的核苷酸掺入中的作用。
DOI：
10.1128/mcb.00851-06
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发表时间：
2006
期刊：
影响因子：
--
通讯作者：
Prakash,Satya
中科院分区：
文献类型：
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作者：
Johnson,RobertE;Haracska,Lajos;Prakash,Louise;Prakash,Satya
人DNA聚合酶I（Pol I）与其他DNA聚合酶的不同之处在于它表现出显著的模板特异性，与相对的嘧啶相比，它是更有效和准确的相对模板嘌呤。具有模板A和引入的dTTP以及具有模板G和引入的dCTP的Pol I的晶体结构已经揭示，在Pol I活性位点中，模板嘌呤采用顺式构象并且与保持反构象的引入的嘧啶形成Hoogsteen碱基对。通过使用2-氨基嘌呤和嘌呤作为模板残基，它们保留了正常的N7位置，但缺少A的N6或G的O6，在这里，我们提供的证据表明，虽然N6处的氢键对于与模板A相对的T的有效掺入是不必要的，但O6处的氢键是与模板G相对的C掺入的先决条件。为了进一步分析O6和N7氢键对Pol i DNA合成的贡献，我们已经检查了它通过6O-甲基鸟嘌呤和8-氧代鸟嘌呤损伤进行复制的能力，这些损伤分别影响模板G的O6和N7位置。我们从这些研究中得出结论，对于与模板A相反的T熟练掺入，仅需要N7氢键，但对于与模板G相反的C熟练掺入，N7和O 6上的氢键都是必需的。与模板A相反，N6氢键对于熟练的T掺入的可免除性具有重要的生物学意义，因为这将赋予Pol i通过损伤复制的能力，这些损伤损害模板A的N1和N6位置处的沃森-克里克氢键潜力。
Human DNA polymerase ι (Pol ι) differs from other DNA polymerases in that it exhibits a marked template specificity, being more efficient and accurate opposite template purines than opposite pyrimidines. The crystal structures of Pol ι with template A and incoming dTTP and with template G and incoming dCTP have revealed that in the Pol ι active site, the templating purine adopts asynconformation and forms a Hoogsteen base pair with the incoming pyrimidine which remains in theanticonformation. By using 2-aminopurine and purine as the templating residues, which retain the normal N7 position but lack the N6of an A or the O6of a G, here we provide evidence that whereas hydrogen bonding at N6is dispensable for the proficient incorporation of a T opposite template A, hydrogen bonding at O6is a prerequisite for C incorporation opposite template G. To further analyze the contributions of O6and N7 hydrogen bonding to DNA synthesis by Pol ι, we have examined its proficiency for replicating through the6O-methyl guanine and 8-oxoguanine lesions, which affect the O6and N7 positions of template G, respectively. We conclude from these studies that for proficient T incorporation opposite template A, only the N7 hydrogen bonding is required, but for proficient C incorporation opposite template G, hydrogen bonding at both the N7 and O6is an imperative. The dispensability of N6hydrogen bonding for proficient T incorporation opposite template A has important biological implications, as that would endow Pol ι with the ability to replicate through lesions which impair the Watson-Crick hydrogen bonding potential at both the N1 and N6positions of templating A.