Folding of a large ribozyme during transcription and the effect of the elongation factor NusA
Folding of a large ribozyme during transcription and the effect of the elongation factor NusA
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DOI：
10.1073/pnas.96.17.9545
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发表时间：
1999-08-17
影响因子：
11.1
通讯作者：
Sosnick, TR
中科院分区：
文献类型：
--
作者：
Pan, T;Artsimovitch, I;Sosnick, TR
我们通过监测其催化活性的外观，比较了枯草芽孢杆菌RNase P的体外转录启动折叠核酶到全长变性状态的再折叠。在37摄氏度时，Mg2+启动的野生型和循环排列核酶的再折叠需要几分钟，并受到动力学陷阱的限制。T7 RNA聚合酶的转录改变了这两种RNA的折叠途径，并引入了新的动力学陷阱。核心大肠杆菌RNA聚合酶的转录产生相同的结果，尽管其延伸率慢了4倍。然而，它的延伸因子NusA的存在使大肠杆菌RNA聚合酶的环状排列核酶的转录启动折叠速度加快了100多倍。NusA的作用可能是由其增强转录暂停引起的，因为NusA不能加速转录启动折叠，使用突变RNA聚合酶，突变RNA聚合酶在核酶合成过程中不能暂停或响应NusA。我们得出结论，转录和其中的特定停顿都可以改变rna折叠途径。
We compared in vitro transcription-initiated folding of the ribozyme from Bacillus subtilis RNase P to refolding from the full-length, denatured state by monitoring the appearance of its catalytic activity. At 37 degrees C, Mg2+-initiated refolding of the wild type and a circularly permutate ribozyme takes minutes and is limited by a kinetic trap. Transcription by T7 RNA polymerase alters the folding pathway of both RNAs and introduces new kinetic traps. Transcription by the core Escherichia coli RNA polymerase yields the same result, in spite of its 4-fold-slower elongation rate. However, the presence of its elongation factor NusA accelerates more than ttl-fold the transcription-initiated folding of the circularly, permutated ribozyme by E.coli RNA polymerase. The effect of NusA likely is caused by its enhancement of transcriptional pausing because NusA did not accelerate transcription-initiated folding using a mutant RNA polymerase that failed to pause or respond to NusA during ribozyme synthesis. We conclude that both transcription and specific pausing therein can alter RNA-folding pathways.