Coordination of multiple enzyme activities by a single PCNA in archaeal Okazaki fragment maturation.

Coordination of multiple enzyme activities by a single PCNA in archaeal Okazaki fragment maturation.
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DOI:
10.1038/emboj.2012.12
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发表时间:
2012-03-21
期刊:
影响因子:
11.4
通讯作者:
Bell, Stephen D.
Bell, Stephen D.
中科院分区:
生物学1区
文献类型:
--
作者:
Beattie, Thomas R.;Bell, Stephen D.

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染色体DNA复制需要一个子链-滞后链-被合成为一系列不连续的RNA引发的冈崎片段,随后必须成熟为一个单一的共价DNA链。在这里,我们描述了冈崎片段成熟的重建在体外使用来自古菌硫磺硫化叶菌的蛋白质。六种蛋白质是偶联DNA合成、RNA引物去除和DNA连接所必需和足够的。PolB1、Fen1和Lig1提供所需的催化活性,其活性的协调依赖于DNA滑动钳、增殖细胞核抗原(PCNA)。S. Solfataricus PCNA是异源三聚体,其中每个亚基对结合PolB1、Fen1或Lig1具有不同的特异性。我们的数据表明,最有效的耦合活动发生时,一个单一的PCNA环组织PolB1,Fen1和Lig1到一个复杂的。
Chromosomal DNA replication requires one daughter strand—the lagging strand—to be synthesised as a series of discontinuous, RNA-primed Okazaki fragments, which must subsequently be matured into a single covalent DNA strand. Here, we describe the reconstitution of Okazaki fragment maturation in vitro using proteins derived from the archaeon Sulfolobus solfataricus. Six proteins are necessary and sufficient for coupled DNA synthesis, RNA primer removal and DNA ligation. PolB1, Fen1 and Lig1 provide the required catalytic activities, with coordination of their activities dependent upon the DNA sliding clamp, proliferating cell nuclear antigen (PCNA). S. solfataricus PCNA is a heterotrimer, with each subunit having a distinct specificity for binding PolB1, Fen1 or Lig1. Our data demonstrate that the most efficient coupling of activities occurs when a single PCNA ring organises PolB1, Fen1 and Lig1 into a complex.
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