Structural equilibria in RNA as revealed by 19F NMR

被引:12
|
作者
Arnold, JRP [1 ]
Fisher, J
机构
[1] Univ Leeds, Sch Biol, Leeds LS2 9JT, W Yorkshire, England
[2] Univ Leeds, Sch Chem, Leeds LS2 9JT, W Yorkshire, England
来源
JOURNAL OF BIOMOLECULAR STRUCTURE & DYNAMICS | 2000年 / 17卷 / 05期
关键词
D O I
10.1080/07391102.2000.10506573
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We have incorporated 5-fluorouridine into several sires within a 19-mer RNA modelled on the translational operator of the MS2 bacteriophage. The F-19 NMR spectra demonstrate the different chemical shifts of helical and loop fluorouridines of the hairpin secondary structure. Addition of salt gives rise to a species in which the loop fluorouridine gains the chemical shift of its helical counterparts, due to the formation of the alternative bi-molecular duplex form. This is supported by UV thermal melting behaviour which becomes highly dependent on the RNA concentration. Distinct F-19 NMR signals for duplex and hairpin forms allow the duplex-hairpin equilibrium constant to be determined under a range of conditions, enabling thermodynamic characterisation and its salt dependence to be determined. Mg2+ also promotes duplex formation, but more strongly than Na+, such that at 25 degrees C, 10 mM MgCl2 has a comparable duplex-promoting effect to 300 mM NaCl. A similar effect is observed with Sr2+, but not Ca2+ or Ba2+. Additional hairpin species are observed in the presence of Na+ as well as Mg2+, Ca2+, Sr2+ and Ba2+ ions. The overall, ensemble average, hairpin conformation is therefore salt-dependent. Electrostatic considerations are thus involved in the balance between different hairpin conformers as well as the duplex-hairpin equilibrium. The data presented here demonstrate that F-19 NMR is a powerful tool for the study of conformational. heterogeneity in RNA, which is particularly important for probing the effects of metal ions on RNA structure. The thermodynamic characterisation of duplex-hairpin equilibria will also be valuable in the development of theoretical models of nucleic acid structure.
引用
收藏
页码:843 / 856
页数:14
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