Fig. 5.

Energy landscape of DNA origami assembly. a In our proposed dynamic model of DNA origami folding, the final fate is dictated by the degree of topological stress experienced at the nucleation sites. Besides the formation of the iso I state, favored by a low level of structural frustration, this mechanism enables to travel an alternative folding pathway leading to a typically less populated iso II state, which becomes competitive for structurally frustrated nucleation sites. b Assembly pathway of domain A in the e(2) design according to the values of activation energy for the cooling and melting of both the iso I and iso II isomers. The energy landscape is representative of a DNA origami assembly with two possible outcomes, connected by a post-assembly reconfiguration that passes through an intermediate open form, as shown by single-molecule force spectroscopy measurements (inset). The energy barriers for folding/unfolding of the iso I species are lower than those of the iso II, resulting in the easier formation and reversibility of the former and large hysteresis of the latter. The lower rate of iso II formation might be due to the entropic penalty and elastic energy cost associated to its folding. Here, the scaffold is entangled into a highly ordered structure constituted by many duplexes shorter than the persistence length of double-stranded DNA. Nevertheless, once the kinetic barrier is passed, the energy gain appears to be large enough to reach an absolute minimum from which the structure cannot easily escape. The energy level associated to the open form has been set arbitrarly and is not matter of this study. Note the two-fold effect of magnesium ions on the energy landscape of connected HJs: increasing Mg²⁺ concentrations lead to a decrease in the iso I/iso II conversion rate (orange arrows, previous studies²⁹) and stabilization of the iso II form (blue arrows, this study)