INTERACTION OF RNA-POLYMERASE WITH LACUV5 PROMOTER DNA DURING MESSENGER-RNA INITIATION AND ELONGATION - FOOTPRINTING, METHYLATION, AND RIFAMPICIN-SENSITIVITY CHANGES ACCOMPANYING TRANSCRIPTION INITIATION
INTERACTION OF RNA-POLYMERASE WITH LACUV5 PROMOTER DNA DURING MESSENGER-RNA INITIATION AND ELONGATION - FOOTPRINTING, METHYLATION, AND RIFAMPICIN-SENSITIVITY CHANGES ACCOMPANYING TRANSCRIPTION INITIATION
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DOI：
10.1016/0022-2836(85)90210-4
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发表时间：
1985-01-01
影响因子：
5.6
通讯作者：
GRALLA, JD
中科院分区：
文献类型：
--
作者：
CARPOUSIS, AJ;GRALLA, JD
酶和化学探针用于跟踪大肠杆菌RNA聚合酶沿着lacUV 5启动子DNA在转录起始期间的移动。RNA聚合酶不会从启动子中逃逸，而是在转录物的初始碱基合成期间保持紧密结合。RNA合成的初始阶段涉及通过RNA聚合酶循环反应重复合成和释放长达10个碱基的RNA链，并且该酶对利福平抑制保持敏感。当形成更长的链时，启动子特异性结合被破坏，并且酶与下游DNA序列形成利福平抗性延伸复合物。这种延伸复合物覆盖了> 1/2的DNA，并且缺乏DNA酶I超敏感位点和碱基特异性接触，这些位点是启动子结合RNA聚合酶的特征。因此，lacUV 5 mRNA的合成是由启动子结合的酶复合物引发的，该酶复合物合成mRNA链中的最初9或10个碱基。因此，当制备10个碱基或稍长的链时，与启动子DNA的接触被不可逆地破坏，亚基丢失，并形成真正的延伸复合物。
Enzymatic and chemical probes were used to follow the movement of Escherichia coli RNA polymerase along lacUV5 promoter DNA during transcription initiation. The RNA polymerase does not escape from the promoter but remains tightly bound during the synthesis of the initial bases of the transcript. This initial phase of RNA synthesis involves the reiterative synthesis and release of RNA chains up to 10 bases long via the RNA polymerase cycling reaction and the enzyme remains sensitive to rifampicin inhibition. When longer chains are made, promoter-specific binding is disrupted and the enzyme forms a rifampicin-resistant elongation complex with downstream DNA sequences. This elongation complex covers > 1/2 as much DNA and lacks the DNase I-hypersensitive sites and the base-specific contacts that characterize promoter-bound RNA polymerase. Thus, lacUV5 mRNA synthesis is primed by a promoter-bound enzyme complex that synthesizes the initial 9 or 10 bases in the mRNA chain. Therefore when a chain of 10 bases, or slightly longer, is made, contacts with promoter DNA are irreversibly disrupted, .sigma. subunit is lost, and a true elongation complex is formed.