Observed instability of T7 RNA polymerase elongation complexes can be dominated by collision-induced "bumping"
Observed instability of T7 RNA polymerase elongation complexes can be dominated by collision-induced "bumping"
复制标题
DOI：
10.1074/jbc.m604369200
复制
发表时间：
2006-08-25
影响因子：
4.8
通讯作者：
Martin, Craig T.
中科院分区：
文献类型：
--
作者：
Zhou, Yi;Martin, Craig T.
T7RNA聚合酶以相对较高的速度拉长RNA，并能取代许多紧密结合的蛋白质-DNA复合体。尽管有这些性质，但对停滞的延长复合体的稳定性的测量表明，寿命比多亚单位RNA聚合酶的寿命短得多。在这项工作中，我们证明了停滞复合体的表面不稳定性实际上是由于拖尾RNA聚合酶(沿同一方向移动)取代停滞复合体的作用。此外，两个聚合酶之间的碰撞引起的不稳定性是位置相关的。当前导复合体距离启动子12个碱基或更少时，第二个聚合酶被阻止与启动子结合。当前导复合体距离启动子12~20bp时，拖尾复合体可以结合并产生流产的转录本，但不能取代第一个复合体，因为它处于不稳定的起始构象。只有当前导复合体距离启动子20个碱基对以上时，第二个聚合酶才能结合、启动和取代前导复合体。
T7 RNA polymerase elongates RNA at a relatively high rate and can displace many tightly bound protein-DNA complexes. Despite these properties, measurements of the stability of stalled elongation complexes have shown lifetimes that are much shorter than those of the multisubunit RNA polymerases. In this work, we demonstrate that the apparent instability of stalled complexes actually arises from the action of trailing RNA polymerases (traveling in the same direction) displacing the stalled complex. Moreover, the instability caused by collision between two polymerases is position dependent. A second polymerase is blocked from promoter binding when a leading complex is stalled 12 bp or less from the promoter. The trailing complex can bind and make abortive transcripts when the leading complex is between 12 and 20 bp from the promoter, but it cannot displace the first complex since it is in a unstable initiation conformation. Only when the leading complex is stalled more than 20 bp away from the promoter can a second polymerase bind, initiate, and displace the leading complex.