  Ribonucleic acid (RNA) is involved in many regulatory and catalytic processes
in the cell. The function of any RNA molecule is intimately related with its
structure. In-line probing experiments provide valuable structural datasets for
a variety of RNAs and are used to characterize conformational changes in
riboswitches. However, the structural determinants that lead to differential
reactivities in unpaired nucleotides have not been investigated yet. In this
work we used a combination of theoretical approaches, i.e., classical molecular
dynamics simulations, multiscale quantum mechanical/molecular mechanical
calculations, and enhanced sampling techniques in order to compute and
interpret the differential reactivity of individual residues in several RNA
motifs including members of the most important GNRA and UNCG tetraloop
families. Simulations on the multi ns timescale are required to converge the
related free-energy landscapes. The results for uGAAAg and cUUCGg tetraloops
and double helices are compared with available data from in-line probing
experiments and show that the introduced technique is able to distinguish
between nucleotides of the uGAAAg tetraloop based on their structural
predispositions towards phosphodiester backbone cleavage. For the cUUCGg
tetraloop, more advanced ab initio calculations would be required. This study
is the first attempt to computationally classify chemical probing experiments
and paves the way for an identification of tertiary structures based on the
measured reactivity of non-reactive nucleotides.
