Mechanism of error-free DNA synthesis across N1-methyl-deoxyadenosine by human DNA polymerase-ι
Mechanism of error-free DNA synthesis across N1-methyl-deoxyadenosine by human DNA polymerase-ι
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DOI：
10.1038/srep43904
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发表时间：
2017-03-08
影响因子：
4.6
通讯作者：
Aggarwal, Aneel K.
中科院分区：
文献类型：
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作者：
Jain, Rinku;Choudhury, Jayati Roy;Aggarwal, Aneel K.
N1-甲基脱氧腺苷(1-mea)是由N1原子上的脱氧腺苷甲基化而成。1-MeA由于不能参与Watson-Crick(W-C)碱基配对，对复制型DNA聚合酶具有阻碍作用。在这里，我们确定人类DNA聚合酶IOTA(POLIOTA)如何促进跨1-MEA的无错误复制。稳态动力学分析表明，POL。在与1-MeA相反的正确核苷酸T与错误核苷酸C的结合效率上类似于100倍。为了了解这种选择性的基础，我们确定了POL的三元结构。绑定到模板1-MEA和传入的dTTP或dCTP。在这两种结构中，模板1-MeA旋转到SYN构象，但与dTTP和dCTP配对不同。因此，虽然dTTP参与了稳定的Hoogsteen碱基与1-MeA的配对，但dCTP无法获得“立足点”，并且在很大程度上是无序的。总而言之，我们的动力学和结构研究表明，波尔。与1-MeA相反，保持对正确和错误的传入核苷酸的区分，以保持基因组的完整性。
N1-methyl-deoxyadenosine (1-MeA) is formed by methylation of deoxyadenosine at the N1 atom. 1-MeA presents a block to replicative DNA polymerases due to its inability to participate in Watson-Crick ( W-C) base pairing. Here we determine how human DNA polymerase-iota (Pol iota) promotes error-free replication across 1-MeA. Steady state kinetic analyses indicate that Pol. is similar to 100 fold more efficient in incorporating the correct nucleotide T versus the incorrect nucleotide C opposite 1-MeA. To understand the basis of this selectivity, we determined ternary structures of Pol. bound to template 1-MeA and incoming dTTP or dCTP. In both structures, template 1-MeA rotates to the syn conformation but pairs differently with dTTP versus dCTP. Thus, whereas dTTP partakes in stable Hoogsteen base pairing with 1-MeA, dCTP fails to gain a "foothold" and is largely disordered. Together, our kinetic and structural studies show how Pol. maintains discrimination between correct and incorrect incoming nucleotide opposite 1-MeA in preserving genome integrity.