Modulating RNA Alignment Using Directional Dynamic Kinks: Application in Determining an Atomic-Resolution Ensemble for a Hairpin using NMR Residual Dipolar Couplings

Approaches
that combine experimental data and computational molecular
dynamics (MD) to determine atomic resolution ensembles of biomolecules
require the measurement of abundant experimental data. NMR residual
dipolar couplings (RDCs) carry rich dynamics information, however,
difficulties in modulating overall alignment of nucleic acids have
limited the ability to fully extract this information. We present
a strategy for modulating RNA alignment that is based on introducing
variable dynamic kinks in terminal helices. With this strategy, we
measured seven sets of RDCs in a cUUCGg apical loop and used this
rich data set to test the accuracy of an 0.8 μs MD simulation
computed using the Amber ff10 force field as well as to determine
an atomic resolution ensemble. The MD-generated ensemble quantitatively
reproduces the measured RDCs, but selection of a sub-ensemble was
required to satisfy the RDCs within error. The largest discrepancies
between the RDC-selected and MD-generated ensembles are observed for
the most flexible loop residues and backbone angles connecting the
loop to the helix, with the RDC-selected ensemble resulting in more
uniform dynamics. Comparison of the RDC-selected ensemble with NMR
spin relaxation data suggests that the dynamics occurs on the ps–ns
time scales as verified by measurements of R1ρ relaxation–dispersion data. The RDC-satisfying
ensemble samples many conformations adopted by the hairpin in crystal
structures indicating that intrinsic plasticity may play important
roles in conformational adaptation. The approach presented here can
be applied to test nucleic acid force fields and to characterize dynamics
in diverse RNA motifs at atomic resolution.