Non-specific binding of Na${}^{+}$ and Mg${}^{2+}$ to RNA determined by force spectroscopy methods

Abstract

RNA duplex stability depends strongly on ionic conditions, and inside cells RNAs are exposed to both monovalent and multivalent ions. Despite recent advances, we do not have general methods to quantitatively account for the effects of monovalent and multivalent ions on RNA stability, and the thermodynamic parameters for secondary structure prediction have only been derived at 1M [Na${}^{+}$]. Here, by mechanically unfolding and folding a 20 bp RNA hairpin using optical tweezers, we study the RNA thermodynamics and kinetics at different monovalent and mixed monovalent/Mg${}^{2+}$ salt conditions. We measure the unfolding and folding rupture forces and apply Kramers theory to extract accurate information about the hairpin free energy landscape under tension at a wide range of ionic conditions. We obtain non-specific corrections for the free energy of formation of the RNA hairpin and measure how the distance of the transition state to the folded state changes with force and ionic strength. We experimentally validate the Tightly Bound Ion model and obtain values for the persistence length of ssRNA. Finally, we test the approximate rule by which the non-specific binding affinity of divalent cations at a given concentration is equivalent to that of monovalent cations taken at 100 fold that concentration for small molecular constructs.

main paper (32 pages, 11 figures, 1 table) + supplementary information (15 pages)

References in corpus (9)

• Verification of the Crooks fluctuation theorem and recovery of RNA folding free energies
• Single-molecule derivation of salt dependent base-pair free energies in DNA
• Mechanical unfolding of RNA hairpins
• Forced-unfolding and force-quench refolding of RNA hairpins
• Thermodynamic and kinetic aspects of RNA pulling experiments
• Improving signal-to-noise resolution in single molecule experiments using molecular constructs with short handles
• Slow nucleic acid unzipping kinetics from sequence-defined barriers
• Force dependent fragility in RNA hairpins
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Cited by in corpus (10)

• A nucleotide-level coarse-grained model of RNA
• Computer Folding of RNA Tetraloops: Identification of Key Force Field Deficiencies
• Unravelling Mg${}^{2+}$-RNA binding with atomistic molecular dynamics
• Directly measuring single molecule heterogeneity using force spectroscopy
• Stem-loop formation drives RNA folding in mechanical unzipping experiments
• Ion assisted structural collapse of a single stranded DNA: a molecular dynamics approach
• Cooperativity-Dependent Folding of Single-Stranded DNA
• Force-Dependent Folding Kinetics of Single Molecules with Multiple Intermediates and Pathways
• Efficient methods for determining folding free energies in single-molecule pulling experiments
• Stacking correlation length in single-stranded DNA