Complexity of Guanine Quadruplex Unfolding Pathways Revealed by Atomistic Pulling Simulations

Guanine quadruplexes (GQs) are non-canonical nucleicacid structuresinvolved in many biological processes. GQs formed in single-strandedregions often need to be unwound by cellular machinery, so their mechanochemicalproperties are important. Here, we performed steered molecular dynamicssimulations of human telomeric GQs to study their unfolding. We examinedfour pulling regimes, including a very slow setup with pulling velocityand force load accessible to high-speed atomic force microscopy. Weidentified multiple factors affecting the unfolding mechanism, i.e.,:(i) the more the direction of force was perpendicular to the GQ channelaxis (determined by GQ topology), the more the base unzipping mechanismhappened, (ii) the more parallel the direction of force was, GQ openingand cross-like GQs were more likely to occur, (iii) strand slippagemechanism was possible for GQs with an all-anti patternin a strand, and (iv) slower pulling velocity led to richer structuraldynamics with sampling of more intermediates and partial refoldingevents. We also identified that a GQ may eventually unfold after aforce drop under forces smaller than those that the GQ withstood beforethe drop. Finally, we found out that different unfolding intermediatescould have very similar chain end-to-end distances, which revealssome limitations of structural interpretations of single-moleculespectroscopic data.

Automated summary

In this study, steered molecular dynamics simulations were performed to investigate the unfolding mechanism of human telomeric GQs. The results showed that the direction of force, pulling velocity, and pulling force all affected the unfolding process. Additionally, it was found that a GQ could continue to unfold after a force drop. Furthermore, the study revealed that different unfolding intermediates could have similar chain end-to-end distances, suggesting limitations in structural interpretations of single-molecule spectroscopic data.