Force field based conformational analysis of RNA structural motifs: GNRA tetraloops and their pyrimidine relatives.

The protocol of conformational analysis applied here to ribonucleotide oligomers combines conformational search in the space of torsion angles and energy minimization using the AMBER4.1 force field with a continuum treatment of electrostatic solute-solvent interactions. RNA fragments with 5'-GGGCGNNAGCCU-3' sequences commonly fold into hairpins with four-membered loops. The combinatorial search for acceptable conformations using the MC-SYM program was restricted to loop nucleotides and yielded roughly 1500 structures being compatible with a double-stranded stem. After energy minimization by the JUMNA program (without applying any experimental constraints), these structures converged into an ensemble of 74 different conformers including 26 structures which contained the sheared G-A base pair observed in experimental studies of GNRA tetraloops. Energetic analysis shows that inclusion of solvent electrostatic effects is critically important for the selection of conformers that agree with experimentally determined structures. The continuum model accounts for solvent polarization by means of the electrostatic reaction field. In the case of GNRA loop sequences, the contributions of the reaction field shift relative stabilities towards conformations showing most of the structural features derived from NMR studies. The agreement of computed conformations with the experimental structures of GAAA, GCAA, and GAGA tetraloops suggests that the continuum treatment of the solvent represents a definitive improvement over methods using simple damping models in electrostatic energy calculations. Application of the procedure described here to the evaluation of the relative stabilities of conformers resulting from searching the conformational space of RNA structural motifs provides some progress in (non-homology based) RNA 3D-structure prediction.

Structural investigations on small RNA molecules using different force field methods.

The structures of two kinds of small RNA hairpin loops were investigated. As an example of four-membered loops the family of GNNA-loops was chosen, and the influence of a variation of the two middle bases on the rather uncommon G-A base pair was examined. Comparison of the resulting structures showed surprisingly little dependence on the sequence for the overall structure and for the G-A base pair geometry. As an example of three-membered loops a systematic investigation on the loop-sequences UUU, AUU, UUA, GUU, UUG was carried out. Here, not only the loop sequence but also the influence of the closing pair was examined. Again the structural variation with different sequences was very small. Both calculations were used to compare the AMBER 4.0 and the JUMNA IV force field programs. Though both programs show a quite different approach to modeling RNA structures the resulting geometries are comparable.