Roles of the Four DNA Polymerases of the Crenarchaeon Sulfolobus solfataricus and Accessory Proteins in DNA Replication
Roles of the Four DNA Polymerases of the Crenarchaeon Sulfolobus solfataricus and Accessory Proteins in DNA Replication
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DOI：
10.1074/jbc.m111.258038
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发表时间：
2011-09-09
影响因子：
4.8
通讯作者：
Guengerich, F. Peter
中科院分区：
文献类型：
--
作者：
Choi, Jeong-Yun;Eoff, Robert L.;Guengerich, F. Peter
极端嗜热泉古菌Sulfolobus solfataricus P2编码三个B家族DNA聚合酶基因B1（Dpo 1）、B2（Dpo 2）和B3（Dpo 3），以及一个Y家族DNA聚合酶基因Dpo 4，它们与真核生物对应物相关。所有四种DNA聚合酶的mRNA和蛋白质在所有生长阶段均组成型表达。Dpo_2和Dpo_3在体外具有很低的DNA聚合酶和3'至5'核酸外切酶活性。稳态动力学效率（k（猫）/K-m）为正确的核苷酸插入DPO 2和DPO 3是几个数量级小于DPO 1和DPO 4。这两个辅助蛋白增殖细胞核抗原和钳加载复制因子C促进DNA合成与DPO 3，与DPO 1和DPO 4，但非常弱与DPO 2。DNA合成DPO 2和DPO 3显着减少单链结合蛋白，相反，DPO 1和DPO 4。在增殖细胞核抗原，复制因子C，和单链结合蛋白的存在下的DNA合成是最进行性的DPO 1，而DNA损伤旁路是最有效的DPO 4。Dpo 2和Dpo 3都绕过了次黄嘌呤和8-氧代鸟嘌呤，但Dpo 1没有。Dpo 2和Dpo 3分别绕过尿嘧啶和顺式环丁烷胸腺嘧啶二聚体。高浓度的Dpo 2或Dpo 3没有减弱Dpo 1或Dpo 4的DNA合成。我们的结论是，DPO 2和DPO 3是功能少得多，更热不稳定性比DPO 1和DPO 4在体外，但有旁路活动的次黄嘌呤，8-氧代鸟嘌呤，尿嘧啶或顺式-顺式环丁烷胸腺嘧啶二聚体，这表明他们的催化作用有限的translesion DNA合成过去脱氨基，氧化的基础病变和/或紫外线诱导的损伤。
The hyperthermophilic crenarchaeon Sulfolobus solfataricus P2 encodes three B-family DNA polymerase genes, B1 (Dpo1), B2 (Dpo2), and B3 (Dpo3), and one Y-family DNA polymerase gene, Dpo4, which are related to eukaryotic counterparts. Both mRNAs and proteins of all four DNA polymerases were constitutively expressed in all growth phases. Dpo2 and Dpo3 possessed very low DNA polymerase and 3' to 5' exonuclease activities in vitro. Steady-state kinetic efficiencies (k(cat)/K-m) for correct nucleotide insertion by Dpo2 and Dpo3 were several orders of magnitude less than Dpo1 and Dpo4. Both the accessory proteins proliferating cell nuclear antigen and the clamp loader replication factor C facilitated DNA synthesis with Dpo3, as with Dpo1 and Dpo4, but very weakly with Dpo2. DNA synthesis by Dpo2 and Dpo3 was remarkably decreased by single-stranded binding protein, in contrast to Dpo1 and Dpo4. DNA synthesis in the presence of proliferating cell nuclear antigen, replication factor C, and single-stranded binding protein was most processive with Dpo1, whereas DNA lesion bypass was most effective with Dpo4. Both Dpo2 and Dpo3, but not Dpo1, bypassed hypoxanthine and 8-oxoguanine. Dpo2 and Dpo3 bypassed uracil and cis-syn cyclobutane thymine dimer, respectively. High concentrations of Dpo2 or Dpo3 did not attenuate DNA synthesis by Dpo1 or Dpo4. We conclude that Dpo2 and Dpo3 are much less functional and more thermolabile than Dpo1 and Dpo4 in vitro but have bypass activities across hypoxanthine, 8-oxoguanine, and either uracil or cis-syn cyclobutane thymine dimer, suggesting their catalytically limited roles in translesion DNA synthesis past deaminated, oxidized base lesions and/or UV-induced damage.