Template strand switching by T7 RNA polymerase
Template strand switching by T7 RNA polymerase
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DOI：
10.1074/jbc.273.17.10253
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发表时间：
1998-04-24
影响因子：
4.8
通讯作者：
McAllister, WT
中科院分区：
文献类型：
--
作者：
Rong, M;Durbin, RK;McAllister, WT
T7 RNA聚合酶（RNAP）能够穿过双链体DNA的模板（T）链中的多种不连续性，包括切口、缺口和分支连接，其中T链的3'端不与非模板（NT）链互补。产物代表T链的忠实拷贝，没有插入或缺失。在具有突出的3'末端的双链模板上，聚合酶能够插入NT链的游离3'末端并将其用作新的T链T7 RNAP绕过T链中断的能力取决于延伸复合物的稳定性，预期稳定局部RNA：DNA杂合体的序列（如T链中C-6区的存在）显著降低RNAP的解离，同时仍允许酶插入新的3'末端。当酶到达模板的末端时（即当合成径流产物时），观察到对RNAP释放的类似影响，导致在多轮转录期间RNA产物的产率显著不同。
T7 RNA polymerase (RNAP) is able to traverse a variety of discontinuities in the template (T) strand of duplex DNA, including nicks, gaps, and branched junctions in which the 3' end of the T strand is not complementary to the non-template (NT) strand. The products represent a faithful copy of the T strand, with no insertions or deletions. On double-stranded templates having protruding 3' ends the polymerase is able to insert the free 3' end of the NT strand and to utilize this as a new T strand ("turn around transcription"), resulting in the anomalous production of high molecular weight transcripts.The capacity of T7 RNAP to bypass interruptions in the T strand depends upon the stability of the elongation complex, Sequences that are expected to stabilize a local RNA:DNA hybrid (such as the presence of a C-6, tract in the T strand) dramatically reduce dissociation of the RNAP while still allowing the enzyme to insert a new 3' end. Similar effects on RNAP release are observed when the enzyme reaches the end of a template (i.e. when synthesizing runoff products), resulting in markedly different yields of RNA product during multiple rounds of transcription.