posted on 1997-05-20, 00:00authored byParag V. Sahasrabudhe, William H. Gmeiner
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.