We make two new contributions to the problem of calculating pseudoknot-free folding pathways with minimum energy barrier between pairs (A, B) of RNA secondary structures. Our first contribution pertains to a problem posed by Morgan and Higgs: find a min-barrier direct folding pathway for a simple energy model in which each base pair contributes −1. In a direct folding pathway, intermediate structures contain only base pairs in A and B and are of length |A△B| (the size of the symmetric difference of the two structures). We show how to solve this problem exactly, using techniques for deconstructing bipartite graphs. The problem is NP-hard and so our algorithm requires exponential time in the worst case but performs quite well empirically on pairs of structures that are hundreds of nucleotides long. Our second contribution shows that for the simple energy model, repeatedly adding or removing a base pair from A ∪ B along a pathway is not useful in minimizing the energy barrier. Two consequences of this result are that (i) the problem of determining the min-barrier pseudoknot-free folding pathway from the space of direct pathways with repeats is NP-hard and (ii) our new algorithm finds the min-barrier pathway not only from the space of direct folding pathways but in fact from the space of direct pathways with repeats.

We present an iterative algorithm, Oxfold, in the framework of stochastic context-free grammars, that emulates the kinetics of RNA folding in a simplified way, in combination with a molecular evolution model. This method improves considerably on existing grammatical models that do not consider folding kinetics. Additionally, the model compares favourably to non-kinetic thermodynamic models.