The Graduate Program in Biochemistry, Graduate Center of the City University of New York, New York, New York 10016, United States.
Biochemistry. 2011 Mar 22;50(11):1875-84. doi: 10.1021/bi101863g. Epub 2011 Feb 4.
The RNA-directed RNA polymerase P2 from cystovirus ϕ6 catalyzes the de novo synthesis of positive and negative strands of the viral double-stranded RNA genome. P2 is mobile on the slow, microsecond to millisecond time scale with various motional modes, putatively assisting in RNA translocation and catalysis. Here we investigate the influence of the extreme 3'-end sequence of the single-stranded RNA templates and the nature of the substrate nucleotide triphosphates on these motional modes using multiple-quantum NMR spectroscopy. We find that P2, in the presence of templates bearing the proper genomic 3'-ends or the preferred initiation nucleotide, displays unique dynamic signatures that are different from those in the presence of nonphysiological templates or substrates. This suggests that dynamics may play a role in the fidelity of recognition of the correct substrates and template sequences to initiate RNA polymerization.
囊状噬菌体 ϕ6 的 RNA 指导的 RNA 聚合酶 P2 催化病毒双链 RNA 基因组的正链和负链的从头合成。P2 在慢的、微秒到毫秒的时间尺度上是移动的，具有各种运动模式，推测有助于 RNA 易位和催化。在这里，我们使用多量子 NMR 光谱法研究了单链 RNA 模板的极端 3'-末端序列和底物核苷酸三磷酸的性质对这些运动模式的影响。我们发现，在存在带有适当基因组 3'-末端或首选起始核苷酸的模板的情况下，P2 显示出独特的动态特征，与存在非生理模板或底物的情况下不同。这表明动力学可能在识别正确的底物和模板序列以启动 RNA 聚合的准确性中发挥作用。
