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Biological macromolecules have evolved to fold and operate in the crowded environment of the cell. We have shown previously that molecular crowding stabilizes folded RNA structures. Here we report SAXS measurements on a 64 kDa bacterial group I ribozyme in the presence of mono- and divalent ions and PEG crowders of different molecular weight. These experiments show that crowders always stabilize the folded RNA, but this stabilization is weaker in NaCl solutions than MgCl2 solutions. Additionally, we find that RNAs with the same global structure, parametrized by Rg, have different scattering functions depending upon the ratio of electrostatic and entropic stabilization by ions and crowders, respectively. We quantify this difference using the scattering length per scattering volume and find that this ratio is larger for RNAs that fold in lower ionic strength solutions due to the higher crowder content. We conclude that lower RNA flexibility, or reduced configurational entropy, widens the free energy gap between the unfolded and folded RNA in crowded MgCl2 solutions.

Crowders Perturb the Entropy of RNA Energy Landscapes to Favor Folding