RNA-Puzzles Round IV: 3D structure predictions of four ribozymes and two aptamers

Abstract

RNA-Puzzles is a collective endeavor dedicated to the advancement and improvement of RNA 3D structure prediction. With agreement from crystallographers, the RNA structures are predicted by various groups before the publication of the crystal structures. We now report the prediction of 3D structures for six RNA sequences: four nucleolytic ribozymes and two riboswitches. Systematic protocols for comparing models and crystal structures are described and analyzed. In these six puzzles, we discuss (i) the comparison between the automated web servers and human experts; (ii) the prediction of coaxial stacking; (iii) the prediction of structural details and ligand binding; (iv) the development of novel prediction methods; and (v) the potential improvements to be made. We show that correct prediction of coaxial stacking and tertiary contacts is essential for the prediction of RNA architecture, while ligand binding modes can only be predicted with low resolution and simultaneous prediction of RNA structure with accurate ligand binding still remains out of reach. All the predicted models are available for the future development of force field parameters and the improvement of comparison and assessment tools.

Keywords: RNA structure; aptamer; modeling; prediction; ribozyme.

FIGURE 1.

The distributions of structure comparison metrics of the six puzzles. The distribution of structure assessment metrics is plotted as the violin plots where each prediction model is shown as a solid line in the violin plot. Thus, the vertical spread displays the range of values for a given metric in all predicted models and the horizontal breadth reflects the number of predicted models at a given value. The assessment metrics include (A) RMSD, (B) Deformation Index (Parisien et al. 2009), (C) Clash score, (D) the Mean of Circular Quantities (Zok et al. 2014) and (E–H)Interaction network fidelity (Parisien et al. 2009) for all parameters together, Watson–Crick pairs, non-Watson–Crick pairs and stacking.

FIGURE 2.

Puzzle 9: 5-hydroxytryptophan riboswitch aptamer. (A) 3D structure comparison between the predicted model with the lowest RMSD (Chen group model 7, shown in blue) and the reference structure in green. (B) The heatmap shows the deformation profile, poorly predicted regions are shown in red; the red regions concentrate on the loop escaping P2 and P3 with the linking region between P2 and P3. (C) The numbering and secondary structure of the reference structure.

FIGURE 3.

Puzzle15: Hammerhead ribozyme. (A) 3D structure comparison between the predicted model with the lowest RMSD (Adamiak model 8, shown in blue) and the reference structure in green. (B) The heatmap shows the deformation profile, in which the poorly predicted regions are shown in red. The largest deviations happen between the 5′- and the 3′-ends. The large deviations, between the 5′- and 3′-ends in the blue box, are artifactual as discussed in the text (see also (D)). (C) When the 5′- and the 3′-ends are removed from evaluation, RNAComposer model automated server model 1 in the round 1 prediction ranks the best. The heatmap shows the deformation profile, in which the poorly predicted regions are shown in red. calculations of the heatmap. (D) The secondary structure of the reference structure. Nucleotides 1–6 and 63–68 (the gray dashes) within the blue rectangle do not form intramolecular base-pairing but instead, they form base pairs with symmetry-related molecules in the crystal (see text for discussion).

FIGURE 4.

Puzzle17: Pistol ribozyme. (A) 3D structure comparison between the predicted model with the lowest RMSD (SimRNA model 1 in the second round prediction, shown in blue) and the reference structure in green. (B) The heatmap shows the deformation profile, in which the poorly predicted regions are shown in red. The deviations are limited to the last two nucleotides of the long strand and the 5′-end of the short second strand (extremities of P3). (C) The secondary structure of the reference structure.

FIGURE 5.

Puzzle19: Twister sister ribozyme 1. (A) 3D structure comparison between the predicted model with the lowest RMSD (Chen group model 1, shown in blue) and the reference structure in green. (B) The heatmap shows the deformation profile, in which the poorly predicted regions are shown in red. (C) The secondary structure of the reference structure.

FIGURE 6.

Puzzle 20: Twister sister ribozyme 2. (A) 3D structure comparison between the predicted model with the lowest RMSD (Bujnicki group model 4, shown in blue) and the reference structure in green. (B) The heatmap shows the deformation profile, in which the poorly predicted regions are shown in red. (C) The secondary structure of the reference structure.

FIGURE 7.

Puzzle 21: Guanidine III Riboswitch. (A) 3D structure comparison between the predicted model with the lowest RMSD (LORES approach from Das group model 1, shown in blue) and the reference structure in green. (B) The heatmap shows the deformation profile, in which the poorly predicted regions are shown in red. The largest deviations occur at the single strand leaving P1 to join P2 (UCAG). (C) The secondary structure of the reference structure. (D) The guanidine binding site in the crystal structure, where guanidine is shown as magenta and proximal contacts are shown as cyan dashed lines.