Lee J, Chastain P D, Kusakabe T, Griffith J D, Richardson C C
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School Boston, Massachusetts 02115, USA.
Mol Cell. 1998 Jun;1(7):1001-10. doi: 10.1016/s1097-2765(00)80100-8.
The coordinated synthesis of both leading and lagging DNA strands is thought to involve a dimeric DNA polymerase and a looping of the lagging strand so that both strands can be synthesized in the same direction. We have constructed a minicircle with a replication fork that permits an assessment of the stoichiometry of the proteins and a measurement of the synthesis of each strand. The replisome consisting of bacteriophage T7 DNA polymerase, helicase, primase, and single-stranded DNA-binding protein mediates coordinated replication. The criteria for coordination are fulfilled: (1) a replication loop is formed, (2) leading and lagging strand synthesis are coupled, (3) the lagging strand polymerase recycles from one Okazaki fragment to another, and (4) the length of Okazaki fragments is regulated. T7 single-stranded DNA-binding protein is essential for coordination.
前导链和后随链DNA的协同合成被认为涉及一种二聚体DNA聚合酶以及后随链的环化，以便两条链都能以相同方向合成。我们构建了一个带有复制叉的微型环，其允许对蛋白质的化学计量进行评估并测量每条链的合成情况。由噬菌体T7 DNA聚合酶、解旋酶、引发酶和单链DNA结合蛋白组成的复制体介导协同复制。协同的标准得以满足：（1）形成了一个复制环；（2）前导链和后随链的合成相互偶联；（3）后随链聚合酶从一个冈崎片段循环至另一个冈崎片段；（4）冈崎片段的长度受到调控。T7单链DNA结合蛋白对于协同作用至关重要。
