RNA and DNA are rapidly emerging
as targets for therapeutics and
as potential frameworks for nanotechnology. Accurate methods for predicting
and designing structures and dynamics of nucleic acids would accelerate
progress in these and other applications. Suitable approximations
for modeling nucleic acids are being developed but require validation
against disparate experimental observations. Here, nuclear magnetic
resonance spectra for RNA and DNA single strands, CAAU and UCAAUC,
are used as benchmarks to test molecular dynamics simulations with
AMBER force fields OL3 and ROC-RNA for RNA and BSC1 for DNA. A detailed
scheme for making comparisons is also presented. The results reflect
recent progress in approximations and reveal remaining challenges.
Nowadays different experimental techniques, such as single molecule or relaxation experiments, can provide dynamic properties of biomolecular systems, but the amount of detail obtainable with these methods is often limited in terms of time or spatial resolution. Here we use state-of-the-art computational techniques, namely atomistic molecular dynamics and Markov state models, to provide insight into the rapid dynamics of short RNA oligonucleotides, in order to elucidate the kinetics of stacking interactions. Analysis of multiple microsecond-long simulations indicates that the main relaxation modes of such molecules can consist of transitions between alternative folded states, rather than between random coils and native structures. After properly removing structures that are artificially stabilized by known inaccuracies of the current RNA AMBER force field, the kinetic properties predicted are consistent with the timescales of previously reported relaxation experiments.
Single-stranded regions of RNA are
important for folding of sequences
into 3D structures and for design of therapeutics targeting RNA. Prediction
of ensembles of 3D structures for single-stranded regions often involves
classical mechanical approximations of interactions defined by quantum
mechanical calculations on small model systems. Nuclear magnetic resonance
(NMR) spectra and molecular dynamics (MD) simulations of short single
strands provide tests for how well the approximations model many of
the interactions. Here, the NMR spectra for UCUCGU at 2, 15, and 30
°C are compared to simulations with the AMBER force fields, OL3
and ROC-RNA. This is the first such comparison to an oligoribonucleotide
containing an internal guanosine nucleotide (G). G is particularly
interesting because of its many H-bonding groups, large dipole moment,
and proclivity for both syn and anti conformations. Results reveal
formation of a G amino to phosphate non-bridging oxygen H-bond. The
results also demonstrate dramatic differences in details of the predicted
structures. The variations emphasize the dependence of predictions
on individual parameters and their balance with the rest of the force
field. The NMR data can serve as a benchmark for future force fields.
We have solved at 1.07 Å resolution the X-ray crystal structure of a polyriboadenylic acid (poly(rA)) parallel and continuous double helix. Fifty-nine years ago, double helices of poly(rA) were first proposed to form at acidic pH. Here, we show that 7-mer oligo(rA), i.e. rA7, hybridizes and overlaps in all registers at pH 3.5 to form stacked double helices that span the crystal. Under these conditions, rA7 forms well-ordered crystals, whereas rA6 forms fragile crystalline-like structures, and rA5, rA8 and rA11 fail to crystallize. Our findings support studies from ∼50 years ago: one showed using spectroscopic methods that duplex formation at pH 4.5 largely starts with rA7 and begins to plateau with rA8; another proposed a so-called ‘staggered zipper’ model in which oligo(rA) strands overlap in multiple registers to extend the helical duplex. While never shown, protonation of adenines at position N1 has been hypothesized to be critical for helix formation. Bond angles in our structure suggest that N1 is protonated on the adenines of every other rAMP−rAMP helix base pair. Our data offer new insights into poly(rA) duplex formation that may be useful in developing a pH sensor.
The Potential, U, is Essential: The many conformations from
a simulation produce a free energy surface. The surface is realistic, if the
force field, FF, is. The database of RNA sequences is exploding, but knowledge
of energetics, structures, and dynamics lags behind. All-atom computational
methods, such as molecular dynamics, hold promise for closing this gap. New
algorithms and faster computers have accelerated progress in improving the
reliability and accuracy of predictions. Currently, the methods can facilitate
refinement of experimentally determined NMR and x-ray structures, but are less
reliable for predictions based only on sequence. Much remains to be discovered,
however, about the many molecular interactions driving RNA folding and the best
way to approximate them quantitatively. The large number of parameters required
means that a wide variety of experimental results will be required to benchmark
force fields and different approaches. As computational methods become more
reliable and accessible, they will be used by an increasing number of
biologists, much as x-ray crystallography has expanded. Thus, many fundamental
physical principles underlying the computational methods are described. This
review presents a summary of the current state of molecular dynamics as applied
to RNA. It is designed to be helpful to students, postdoctoral fellows, and
faculty who are considering or starting computational studies of RNA.
Geometrical restraints provide key structural information for the determination of biomolecular structures at lower resolution by experimental methods such as crystallography or cryo-electron microscopy. In this work, restraint targets for nucleic acids bases are derived from three different sources and compared: small-molecule crystal structures in the Cambridge Structural Database (CSD), ultrahigh-resolution structures in the Protein Data Bank (PDB) and quantum-mechanical (QM) calculations. The best parameters are those based on CSD structures. After over two decades, the standard library of Parkinson et al. [(1996), Acta Cryst. D52, 57–64] is still valid, but improvements are possible with the use of the current CSD database. The CSD-derived geometry is fully compatible with Watson–Crick base pairs, as comparisons with QM results for isolated and paired bases clearly show that the CSD targets closely correspond to proper base pairing. While the QM results are capable of distinguishing between single and paired bases, their level of accuracy is, on average, nearly two times lower than for the CSD-derived targets when gauged by root-mean-square deviations from ultrahigh-resolution structures in the PDB. Nevertheless, the accuracy of QM results appears sufficient to provide stereochemical targets for synthetic base pairs where no reliable experimental structural information is available. To enable future tests for this approach, QM calculations are provided for isocytosine, isoguanine and the iCiG base pair.
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