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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Minichromosome maintenance helicase activity is controlled by N- and C-terminal motifs and requires the ATPase domain helix-2 insert
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Minichromosome maintenance helicase activity is controlled by N- and C-terminal motifs and requires the ATPase domain helix-2 insert

机译：微型染色体维持解旋酶活性受N和C端基序控制，需要ATPase域helix-2插入

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摘要

The minichromosome maintenance (MCM) proteins are essential conserved proteins required for DNA replication in archaea and eukaryotes. MCM proteins are believed to provide the replicative helicase activity that unwinds template DNA ahead of the replication fork. Consistent with this hypothesis, MCM proteins can form hexameric complexes that possess ATP-dependent DNA unwinding activity. The molecular mechanism by which the energy of ATP hydrolysis is harnessed to DNA unwinding is unknown, although the ATPase activity has been attributed to a highly conserved AAA+ family ATPase domain. Here we show that changes to Nand C-terminal motifs in the single MCM protein from the archaeon Methanothermobacter thermautotrophicus (MthMCM) can modulate ATP hydrolysis, DNA binding, and duplex unwinding. Furthermore, these motifs appear to influence the movement of the beta-alpha-beta insert in helix-2 of the MCM ATPase domain. Removal of this motif from MthMCM increased dsDNA-stimulated ATIP hydrolysis and increased the affinity of the mutant complex for ssDNA and dsDNA. Deletion of the helix-2 insert additionally resulted in the abrogation of DNA unwinding. Our results provide significant insight into the molecular mechanisms used by the MCM helicase to both regulate and execute DNA unwinding.
机译：微染色体维持（MCM）蛋白是古细菌和真核生物中DNA复制所需的必不可少的保守蛋白。据信MCM蛋白具有复制解旋酶活性，可在复制叉之前解开模板DNA。与此假设一致，MCM蛋白可以形成具有ATP依赖的DNA解链活性的六聚体复合物。尽管将ATPase活性归因于高度保守的AAA +家族ATPase结构域，但尚不知道将ATP水解的能量用于DNA解旋的分子机制。在这里，我们显示从古细菌甲烷嗜热杆菌（MthMCM）到单个MCM蛋白中Nand C端基序的变化可以调节ATP水解，DNA结合和双链解绕。此外，这些基序似乎会影响MCM ATPase域螺旋2中的beta-alpha-beta插入片段的运动。从MthMCM中除去此基序会增加dsDNA刺激的ATIP水解，并增加突变复合物对ssDNA和dsDNA的亲和力。删除螺旋2插入物还导致废除DNA解链。我们的结果为MCM解旋酶用于调控和解链DNA的分子机制提供了重要的见识。

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