Structural insight into the DNA polymerase β deoxyribose phosphate lyase mechanism
Structural insight into the DNA polymerase β deoxyribose phosphate lyase mechanism
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DOI：
10.1016/j.dnarep.2005.08.009
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发表时间：
2005-12-08
期刊：
影响因子：
3.8
通讯作者：
Wilson, SH
中科院分区：
文献类型：
--
作者：
Prasad, R;Batra, VK;Wilson, SH
大量的生物化学和遗传学研究已经证明DNA聚合酶β（Pol β）参与哺乳动物的碱基切除修复（BER）。Pot β通过促进缺口填充DNA合成和裂解酶从切割的脱碱基位点去除5 '-脱氧核糖磷酸（dRP）基团而参与BER子途径。为了在原子水平上更好地理解dRP裂解酶反应的机制，我们确定了在有切口的DNA中与5 '-磷酸化脱碱基糖类似物复合的Pot β的晶体结构。该DNA配体代表潜在的BER中间体。晶体结构表明，dRP基团结合在非催化结合位点。dRP裂解酶反应中的催化亲核试剂Lys 72和所有其他潜在的二级亲核试剂太远而不能参与对糖的C1'的亲核攻击。预期催化结合位点中dRP基团的近似模型表明，需要围绕dRP 3 '-磷酸旋转120度以将ε-氨基Lys 72定位在dRPC 1'附近。该模型还表明，其他几个侧链的位置，以促进β-消除反应。从dRP裂解酶活性位点中的关键残基的突变分析结果来看，似乎底物dRP可以稳定在所观察到的非催化结合构象中，从而阻碍dRP裂解酶活性。由爱思唯尔公司出版
A large number of biochemical and genetic studies have demonstrated the involvement of DNA polymerase beta (Pol beta) in mammalian base excision repair (BER). Pot beta participates in BER sub-pathways by contributing gap filling DNA synthesis and lyase removal of the 5'-deoxyribose phosphate (dRP) group from the cleaved abasic site. To better understand the mechanism of the dRP lyase reaction at an atomic level, we determined a crystal structure of Pot beta complexed with 5'-phosphorylated abasic sugar analogs in nicked DNA. This DNA ligand represents a potential BER intermediate. The crystal structure reveals that the dRP group is bound in a non-catalytic binding site. The catalytic nucleophile in the dRP lyase reaction, Lys72, and all other potential secondary nucleophiles, are too far away to participate in nucleophilic attack on the C1' of the sugar. An approximate model of the dRP group in the expected catalytic binding site suggests that a rotation of 120 degrees about the dRP 3'-phosphate is required to position the epsilon-amino Lys72 close to the dRPC1'. This model also suggests that several other side chains are in position to facilitate the beta-elimination reaction. From results of mutational analysis of key residues in the dRP lyase active site, it appears that the substrate dRP can be stabilized in the observed non-catalytic binding conformation, hindering dRP lyase activity. Published by Elsevier B.V.