Calcium is a cofactor of polymerization but inhibits pyrophosphorolysis by the Sulfolobus solfataricus DNA polymerase Dpo4
Calcium is a cofactor of polymerization but inhibits pyrophosphorolysis by the Sulfolobus solfataricus DNA polymerase Dpo4
复制标题
DOI：
10.1021/bi052511
复制
发表时间：
2006-05-16
期刊：
影响因子：
2.9
通讯作者：
Egli, Martin
中科院分区：
文献类型：
--
作者：
Irimia, Adriana;Zang, Hong;Egli, Martin
来自硫磺硫化叶菌的Y-家族DNA聚合酶IV（Dpo 4）作为真核细胞跨损伤聚合酶的模型系统，其与天然和加合DNA的复合物的三维结构已被相当详细地分析。dpo 4缺乏复制型聚合酶中常见的校正核酸外切酶活性，但利用焦磷酸解作用来降低掺入不正确碱基的可能性。Mg 2+是聚合酶和焦磷酸解活性的辅因子。尽管Dpo 4的所有晶体结构都是在Ca 2+存在下获得的，但迄今为止还没有研究用Ca 2+取代Mg 2+对Dpo 4活性的影响。我们在这里表明，Ca 2+（但不是Ba 2+，Co 2+，Cu 2+，Ni 2+，或Zn 2+）是一个辅因子DPO 4催化聚合与天然和8- oxoG-含有DNA模板。在Mg 2+或Ca 2+存在下，dNTP和ddNTP都是聚合酶的底物。相反，没有焦磷酸解发生在Ca 2+的存在下，虽然在活性位点的两个催化金属离子的位置似乎是非常相似的混合Mg 2 +/Ca 2 +-和Ca 2 +-形式的DPO 4晶体。
Y- Family DNA polymerase IV ( Dpo4) from Sulfolobus solfataricus serves as a model system for eukaryotic translesion polymerases, and three- dimensional structures of its complexes with native and adducted DNA have been analyzed in considerable detail. Dpo4 lacks a proofreading exonuclease activity common in replicative polymerases but uses pyrophosphorolysis to reduce the likelihood of incorporation of an incorrect base. Mg2+ is a cofactor for both the polymerase and pyrophosphorolysis activities. Despite the fact that all crystal structures of Dpo4 have been obtained in the presence of Ca2+, the consequences of replacing Mg2+ with Ca2+ for Dpo4 activity have not been investigated to date. We show here that Ca2+ ( but not Ba2+, Co2+, Cu2+, Ni2+, or Zn2+) is a cofactor for Dpo4- catalyzed polymerization with both native and 8- oxoG- containing DNA templates. Both dNTP and ddNTP are substrates of the polymerase in the presence of either Mg2+ or Ca2+. Conversely, no pyrophosphorolysis occurs in the presence of Ca2+, although the positions of the two catalytic metal ions at the active site appear to be very similar in mixed Mg2+/ Ca2+- and Ca2+- form Dpo4 crystals.