  We introduce a method for predicting RNA folding pathways, with an
application to the most important RNA tetraloops. The method is based on the
idea that ensembles of three-dimensional fragments extracted from
high-resolution crystal structures are heterogeneous enough to describe
metastable as well as intermediate states. These ensembles are first validated
by performing a quantitative comparison against available solution NMR data of
a set of RNA tetranucleotides. Notably, the agreement is better with respect to
the one obtained by comparing NMR with extensive all-atom molecular dynamics
simulations. We then propose a procedure based on diffusion maps and Markov
models that makes it possible to obtain reaction pathways and their relative
probabilities from fragment ensembles. This approach is applied to study the
helix-to-loop folding pathway of all the tetraloops from the GNRA and UNCG
families. The results give detailed insights into the folding mechanism that
are compatible with available experimental data and clarify the role of
intermediate states observed in previous simulation studies. The method is
computationally inexpensive and can be used to study arbitrary conformational
transitions.
