The fidelity of DNA polymerase β during distributive and processive DNA synthesis
The fidelity of DNA polymerase β during distributive and processive DNA synthesis
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DOI：
10.1074/jbc.274.6.3642
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发表时间：
1999-02-05
影响因子：
4.8
通讯作者：
Kunkel, TA
中科院分区：
文献类型：
--
作者：
Osheroff, WP;Jung, HK;Kunkel, TA
在碱基切除修复过程中，DNA聚合酶β连续填补1-6个核苷酸的缺口，反映其8-kDa和31-kDa结构域对DNA结合的贡献。在这里，我们报告了在合成过程中，为填补1、5、6或>；300个核苷酸的空白而产生的POLβ的保真度。具有390个碱基缺口的重组大鼠和人聚合酶(Pol)β在分布合成过程中的错误率彼此相似，并且与从组织中纯化的olβ之前的值相似。当连续填充5个核苷酸缺口时，人olβ的碱基替换保真度与361个核苷酸缺口的碱基替换保真度相似，但“紧密”替换的产生速度至少比分布合成高60倍，填充1个核苷酸缺口时的碱基替换保真度高于填充5个核苷酸缺口时的碱基替换保真度，这表明8-kDa结构域对dNTP结合口袋的贡献和/或olβ造成的碱基堆积或DNA结构的差异。然而，1-核苷酸缺口填充是不准确的，甚至会产生复杂的替代-加成错误。最后，在过程合成过程中填补6个核苷酸缺口的单碱基缺失错误率与分布合成填补390个核苷酸缺口时的错误率是无法区分的。因此，polβ的加工性机制不允许酶抑制模板错位。
During base excision repair, DNA polymerase beta fills 1-6-nucleotide gaps processively, reflecting a contribution of both its 8- and 31-kDa domains to DNA binding. Here we report the fidelity of pol beta during synthesis to fill gaps of 1, 5, 6, or >300 nucleotides. Error rates during distributive synthesis by recombinant rat and human polymerase (pol) beta with a 390-base gap are similar to each other and to previous values with pol beta purified from tissues. The base substitution fidelity of human pol beta when processively filling a 5-nucleotide gap is similar to that with a 361-nucleotide gap, but "closely-spaced" substitutions are produced at a rate at least 60-fold higher than for distributive synthesis, Base substitution fidelity when filling a 1-nucleotide gap is higher than when filling a 5-nucleotide gap, suggesting a contribution of the 8-kDa domain to the dNTP binding pocket and/or a difference in base stacking or DNA structure imposed by pol beta. Nonetheless, 1-nucleotide gap filling is inaccurate, even generating complex substitution-addition errors. Finally, the single-base deletion error rate during processive synthesis to fill a 6-nucleotide gap is indistinguishable from that of distributive synthesis to fill a 390-nucleotide gap. Thus the mechanism of processivity by pol beta does not allow the enzyme to suppress template misalignments.