Stability of End-to-End Base Stacking Interactions in Highly Concentrated DNA Solutions

Abstract

At sufficiently high concentrations, gapped DNA (GDNA) constructs - two duplex arms connected by a flexible single stranded linker - form layered (smectic) liquid crystalline (LC) phases, whose nature and stability depend on both end-to-end stacking and side-by-side electrostatic interactions between the duplex arms. We performed synchrotron small angle x-ray scattering (SAXS) experiments over 5-65 C on GDNA constructs that differ only by the terminal base-pairs at the blunt ends of the duplex arms. Two smectic phases are generally observed at the DNA concentrations studied: a bilayer phase at lower temperature, where the duplexes are stacked end-to-end and also positionally ordered within the layers (smectic-B phase), and a monolayer phase at higher temperature with single duplex layer spacing and short-range order within layers (smectic-A phase). Our key finding is that the stability of the bilayer phase varies remarkably among constructs that differ only in their terminal base-pairs. Bilayers formed by constructs with GC termination at both blunt duplex ends melt into the monolayer phase at temperatures up to 30 C higher than for those with AT termination, while mixed (AT/GC) terminations have intermediate stability. Correlating this compositional variation with macroscopic effects on the thermal stability of the bilayer smectic phase enables us to determine the relative strength of base pair specific, end-to-end stacking interactions. Modeling the bilayer melting in terms of a temperature-dependent reduction in the average fraction of end-to-end paired duplexes enables us to estimate stacking free energies in solutions of duplexes at physiologically relevant DNA concentrations.