Solution structures of 5-fluorouracil-substituted RNA duplexes containing G-U wobble base pairs.
The structures and stabilities of three RNA duplexes that differed only in the position of 5-fluorouridine (FUrd) substitution were elucidated using NMR spectroscopy and UV hyperchromicity studies to determine if FUrd substitution altered the structure or stability of RNA duplexes that contained G-U base pairs. The duplexes investigated corresponded to the region of the U4-U6 snRNA complex that contained the 5' terminus of U4 snRNA. The control duplex contained a G-U wobble base pair and also a G-A mismatched base pair. FUrd was substituted in one duplex at the G-U wobble base pair and in the second duplex at an A-U base pair adjacent to the wobble base pair. FUrd substitution slightly destabilized the duplex that contained a G-FU base pair but stabilized the duplex that contained an A-FU base pair. NOESY spectra were used to determine interproton distances, and these distance constraints were used in a restrained molecular dynamics protocol to determine the three-dimensional structures of these RNA duplexes. Analyses of helical parameters, backbone torsion angles, and rms deviations between the final structures revealed no systematic differences due to FUrd substitution in RNA duplexes that contained G-U base pairs. The G-FU base pair adopted wobble geometry, while the G-A mismatch formed a sheared base pair. NOESY spectra in H2O solution revealed the imino 1H from FUrd exchanged more rapidly with solvent than did the Urd imino 1H but did not show the G-FU base pair adopted an ionized structure. Reduced stacking occurred for the G-FU base pair relative to the G-U base pair in the time-averaged structure, and this, rather than ionization of the base pair, was responsible for the slight destabilization of the duplex that contained the G-FU base pair.
Sahasrabudhe
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PV
Eppley Institute for Cancer Research and Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha 68198-6805, USA.
Gmeiner
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D000964:Antimetabolites, Antineoplastic / Q000737:chemistry*; D005472:Fluorouracil / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D009038:Motion; D009690:Nucleic Acid Conformation; D011522:Protons; D012330:RNA, Double-Stranded / Q000737:chemistry*; D012342:RNA, Small Nuclear / Q000737:chemistry*; D015203:Reproducibility of Results; D013816:Thermodynamics
D000964:Antimetabolites, Antineoplastic; D011522:Protons; D012330:RNA, Double-Stranded; D012342:RNA, Small Nuclear; D005472:Fluorouracil