Morrison Paul F, Chattoraj Dhruba K
Division of Bioengineering and Physical Science, ORS, National Institutes of Health, Bethesda, MD 20892-5766, USA.
Plasmid. 2004 Jul;52(1):13-30. doi: 10.1016/j.plasmid.2004.04.001.
For stability, the replication of unit-copy plasmids ought to occur by a highly controlled process. We have characterized the replication dynamics of a unit-copy plasmid F by a replication rate function defined as the probability per unit age interval of the cell cycle that a plasmid will initiate replication. Analysis of baby-machine data [J. Bacteriol. 170 (1988) 1380; J. Bacteriol. 179 (1997) 1393] by stochastics that make no detailed reference to underlying mechanism revealed that this rate function increased monotonically over the cell cycle with rapid increase near cell division. This feature is highly suggestive of a replication control mechanism that is designed to force most plasmids to replicate before cells undergo division. The replication rate function is developed anew from a mechanistic model incorporating the hypotheses that initiators are limiting and that steric hindrance of origins by handcuffing control initiation of replication. The model is based on correctly folded initiator protein monomers arising from an inactive dimer pool via chaperones in limiting amounts, their random distribution to high affinity sites (iterons) at the origin (ori) and an outside locus (incC), the statistical mechanics of bound monomer participation in pairing the two loci (cis-handcuffing), and initiation probability as proportional to the number of non-handcuffed ori-saturated plasmids. Provided cis-handcuffing is present, this model closely accounts for the shape of the replication rate function derived from experiment, and reproduces the observation that replication occurs throughout the cell cycle. Present concepts of iteron-based molecular mechanisms thus appear capable of yielding a quantitative description of unit-copy-number plasmid replication dynamics.
为了保证稳定性，单位拷贝质粒的复制应该通过一个高度受控的过程来进行。我们通过定义为细胞周期中单位年龄间隔内质粒启动复制的概率的复制速率函数，对单位拷贝质粒F的复制动态进行了表征。通过不详细参考潜在机制的随机分析“婴儿机器”数据[《细菌学杂志》170 (1988) 1380；《细菌学杂志》179 (1997) 1393]发现，该速率函数在细胞周期中单调增加，在细胞分裂附近迅速上升。这一特征强烈暗示了一种复制控制机制，其设计目的是迫使大多数质粒在细胞分裂之前进行复制。复制速率函数是从一个机制模型重新推导出来的，该模型包含以下假设：引发剂是有限的，并且通过“手铐”作用对复制起点的空间位阻控制复制的起始。该模型基于以下过程：通过有限数量的伴侣蛋白从无活性二聚体库中产生正确折叠的引发剂蛋白单体，它们随机分布到复制起点（ori）和外部位点（incC）的高亲和力位点（迭代子），结合的单体参与两个位点配对（顺式“手铐”作用）的统计力学，以及起始概率与未被“手铐”束缚且ori饱和的质粒数量成正比。如果存在顺式“手铐”作用，该模型能够很好地解释从实验中得出的复制速率函数的形状，并重现整个细胞周期中都发生复制的观察结果。因此，基于迭代子的分子机制的当前概念似乎能够对单位拷贝数质粒的复制动态进行定量描述。
