Unusual mechanical stability of a minimal RNA kissing complex

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Abstract
By using optical tweezers, we have investigated the mechanical unfolding of a minimal kissing complex with only two G.C base pairs. The loop-loop interaction is exceptionally stable; it is disrupted at forces ranging from 7 to 30 pN, as compared with 14-20 pN for unfolding hairpins of 7 and 11 bp. By monitoring unfolding/folding trajectories of single molecules, we resolved the intermediates, measured their rate constants, and pinpointed the rate-limiting steps. The two hairpins unfold only after breaking the intramolecular kissing interaction, and the kissing interaction forms only after the folding of the hairpins. At forces that favor the unfolding of the hairpins, the entire RNA structure is kinetically stabilized by the kissing interaction, and extra work is required to unfold the metastable hairpins. The strong mechanical stability of even a minimal kissing complex indicates the importance of such loop-loop interactions in initiating and stabilizing RNA dimers in retroviruses.