Beuning P J, Tessmer M R, Baumann C G, Kallick D A, Musier-Forsyth K
Department of Chemistry, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Anal Biochem. 1999 Sep 10;273(2):284-90. doi: 10.1006/abio.1999.4223.
In this study, we use native polyacrylamide gel electrophoresis and one-dimensional NMR spectroscopy to analyze small RNA hairpins containing a UUCG tetraloop. The aggregation state of one RNA 16-mer (5'-CGGCUUCGGUCGACCA-3') in the presence of Mg(2+) was confirmed by laser light scattering. Although it is widely known in the RNA field that some RNAs tend to aggregate, especially when present at high concentrations, the sequence elements responsible for this effect are rarely identified. In this work, we show that Mg(2+)-induced aggregation of the 16-mer RNA hairpin is sensitive to the presence of the 3'-terminal base and a specific 2'-hydroxyl group. Our study highlights the fact that even small changes in a particular RNA sequence can increase its tendency to undergo Mg(2+)-dependent aggregation in an unpredictable manner. Our analysis also shows that native gel electrophoresis is a sensitive probe of RNA conformation with the capability to detect differences apparently caused by subtle base stacking effects at the ends of helices.
在本研究中，我们使用天然聚丙烯酰胺凝胶电泳和一维核磁共振光谱来分析含有UUCG四环的小RNA发夹结构。通过激光光散射确定了一种RNA 16聚体（5'-CGGCUUCGGUCGACCA-3'）在Mg(2+)存在下的聚集状态。尽管在RNA领域广为人知的是，一些RNA倾向于聚集，尤其是在高浓度存在时，但导致这种效应的序列元件却很少被确定。在这项工作中，我们表明Mg(2+)诱导的16聚体RNA发夹的聚集对3'-末端碱基和特定的2'-羟基的存在敏感。我们 的研究突出了这样一个事实，即即使特定RNA序列中的微小变化也可能以不可预测的方式增加其发生Mg(2+)依赖性聚集的倾向。我们的分析还表明，天然凝胶电泳是RNA构象的灵敏探针，能够检测由螺旋末端微妙的碱基堆积效应明显引起的差异。
