title: The essential role of stacking adenines in a two-base-pair RNA kissing complex.
abstract: In minimal RNA kissing complexes formed between hairpins with cognate GACG tetraloops, the two tertiary GC pairs are likely stabilized by the stacking of 5'-unpaired adenines at each end of the short helix. To test this hypothesis, we mutated the flanking adenines to various nucleosides and examined their effects on the kissing interaction. Electrospray ionization mass spectrometry was used to detect kissing dimers in a multiequilibria mixture, whereas optical tweezers were applied to monitor the (un)folding trajectories of single RNA molecules. The experimental findings were rationalized by molecular dynamics simulations. Together, the results showed that the stacked adenines are indispensable for the tertiary interaction. By shielding the tertiary base pairs from solvent and reducing their fraying, the stacked adenines made terminal pairs act more like interior base pairs. The purine double-ring of adenine was essential for effective stacking, whereas additional functional groups modulated the stabilizing effects through varying hydrophobic and electrostatic forces. Furthermore, formation of the kissing complex was dominated by base pairing, whereas its dissociation was significantly influenced by the flanking bases. Together, these findings indicate that unpaired flanking nucleotides play essential roles in the formation of otherwise unstable two-base-pair RNA tertiary interactions.
authors.affiliation: Nanoscale Engineering Graduate Program, University at Albany, State University of New York, 1400 Washington Avenue, Albany, New York 12222, USA.
authors.forename: William
authors.initials: W
authors.lastname: Stephenson
authors.forename: Papa Nii
authors.initials: PN
authors.lastname: Asare-Okai
authors.forename: Alan A
authors.initials: AA
authors.lastname: Chen
authors.forename: Sean
authors.initials: S
authors.lastname: Keller
authors.forename: Rachel
authors.initials: R
authors.lastname: Santiago
authors.forename: Scott A
authors.initials: SA
authors.lastname: Tenenbaum
authors.forename: Angel E
authors.initials: AE
authors.lastname: Garcia
authors.forename: Daniele
authors.initials: D
authors.lastname: Fabris
authors.forename: Pan T X
authors.initials: PT
authors.lastname: Li
mesh_terms: D000225:Adenine; D020029:Base Pairing; D001483:Base Sequence; D007700:Kinetics; D056004:Molecular Dynamics Simulation; D009711:Nucleotides / Q000737:chemistry; D052898:Optical Tweezers; D012313:RNA / Q000737:chemistry* / Q000235:genetics; D013816:Thermodynamics
chemical_list: D009711:Nucleotides; D012313:RNA; D000225:Adenine