On-off regulation of 3' exonuclease excision to DNA polymerization by Exo+ polymerase.
On-off regulation of 3' exonuclease excision to DNA polymerization by Exo+ polymerase.
复制标题
DOI：
10.5483/bmbrep.2003.36.6.525
复制
发表时间：
2003-11
期刊：
影响因子：
--
通讯作者：
Jia Zhang;Kai Li
中科院分区：
文献类型：
--
作者：
Jia Zhang;Kai Li
用3‘端具有核酸外切酶可消化和不能外切核酸酶的3’等位基因特异性引物，对3‘端核酸外切酶在DNA聚合中的作用进行了评价。用外切酶可消化的未修改的3‘端不匹配的引物，外源聚合酶产生依赖于模板的产物。使用3‘端错配的外切酶抗性引物，未发现外源聚合酶扩增产物。作为对照，没有校对活性的聚合酶从3‘错配的引物中产生了依赖于引物的产物。这些数据表明，通过Exo+聚合酶从3‘错配的引物中成功地去除错配是DNA聚合所必需的。除了众所周知的这种错配去除的校对之外，由于未能有效地去除错配的核苷酸，DNA聚合中的提前终止在维持外源DNA复制的高保真度方面起到了关闭开关的作用。
The role of 3' exonuclease excision in DNA polymerization was evaluated in primer extensions using 3' allele-specific primers that had exonuclease-digestible and exonuclease-resistant 3' termini. With exonuclease-digestible unmodified 3' mismatched primers, the exo+ polymerase yielded template-dependent products. Using exonuclease-resistant 3' mismatched primers, no primer-extended product resulted from exo+ polymerase. As a control, polymerase without proofreading activity yielded primer-dependent products from 3' mismatched primers. These data indicated that a successful removal of the mismatch is required for DNA polymerization from the 3' mismatched primers by exo+ polymerase. In addition to the well-known proofreading from this mismatch removal, the premature termination in DNA polymerization, due to the failure of the efficient removal of the mismatched nucleotides, worked as an off-switch in maintaining the high fidelity in DNA replication from exo+ polymerase.