Sequence-dependent conformational differences of small RNAs revealed by native gel electrophoresis

In this study, we use native polyacrylamide gel electrophoresis and one-dim ensional NMR spectroscopy to analyze small RNA hairpins containing a UUCG t etraloop, The aggregation state of one RNA 16-mer (5'-CGGCUUCGGUCGACCA-3') in the presence of Mg2+ was confirmed by laser light scattering. Although i t is widely known in the RNA field that some RNAs tend to aggregate, especi ally when present at high concentrations, the sequence elements responsible for this effect are rarely identified. In this work, we show that Mg2+-ind uced 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 Mg2+-dependent aggregation in an unpredictable man ner. Our analysis also shows that native gel electrophoresis isa sensitive probe of RNA conformation with the capability to detect differences appare ntly caused by subtle base stacking effects at the ends of helices. (C) 199 9 Academic Press.