Structures and operating principles of the replisome
Structures and operating principles of the replisome
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DOI：
10.1126/science.aav7003
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发表时间：
2019-02-22
期刊：
影响因子：
56.9
通讯作者：
Yang, Wei
中科院分区：
文献类型：
--
作者：
Gao, Yang;Cui, Yanxiang;Yang, Wei
在复制叉处协同进行前导链和滞后链DNA合成的复制体的原子细节可视化尚未见报道。我们以噬菌体T7作为模型系统，确定了分辨率高达3.2埃的解旋酶沿DNA移动以及解旋酶 - 聚合酶 - 引发酶复合物参与两条DNA链合成的冷冻电镜结构。螺旋状六聚体解旋酶的每个结构域沿着单链DNA螺旋依次交替移动，类似于AAA + ATP酶（腺苷三磷酸酶）展开肽链的方式。两个滞后链聚合酶连接在引发酶上，准备串联进行冈崎片段合成。来自前导链聚合酶的一个β发夹结构将两条亲代DNA链分离成一个T形叉，从而使紧密偶联的解旋酶能够垂直于下游DNA双链体前进。这些结构揭示了复制体的分子组织和运行原理。
Visualization in atomic detail of the replisome that performs concerted leading- and lagging-DNA strand synthesis at a replication fork has not been reported. Using bacteriophage T7 as a model system, we determined cryo-electron microscopy structures up to 3.2-angstroms resolution of helicase translocating along DNA and of helicase-polymerase-primase complexes engaging in synthesis of both DNA strands. Each domain of the spiral-shaped hexameric helicase translocates sequentially hand-over-hand along a single-stranded DNA coil, akin to the way AAA+ ATPases (adenosine triphosphatases) unfold peptides. Two lagging-strand polymerases are attached to the primase, ready for Okazaki fragment synthesis in tandem. A beta hairpin from the leading-strand polymerase separates two parental DNA strands into a T-shaped fork, thus enabling the closely coupled helicase to advance perpendicular to the downstream DNA duplex. These structures reveal the molecular organization and operating principles of a replisome.