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RNA often folds hierarchically, so that its sequence defines its secondary structure (helical base-paired regions connected by single-stranded junctions), which subsequently defines its tertiary fold. To preserve base-pairing and chain connectivity, the three-dimensional conformations that RNA can explore are strongly confined compared to when secondary structure constraints are not enforced. Using three examples, we studied how secondary structure confines and dictates an RNA's preferred conformations. We made use of Macromolecular Conformations by SYMbolic programming (MC-Sym) fragment assembly to generate RNA conformations constrained by secondary structure. Then, to understand the correlations between different helix placements and orientations, we robustly clustered all RNA conformations by employing unique methods to remove outliers and estimate the best number of conformational clusters. We observed that the preferred conformation (as judged by largest cluster size) for each type of RNA junction molecule tested is consistent with its biological function. Further, the improved quality of models in our pruned datasets facilitates subsequent discrimination using scoring functions based either on statistical analysis (knowledge based) or experimental data.

Clustering to identify RNA conformations constrained by secondary structure