Hidden Complexity in the Isomerization Dynamics of Holliday Junctions

TL;DR

This study reveals that Holliday Junctions exhibit non-ergodic isomerization dynamics with persistent heterogeneity, partitioned into kinetically disconnected clusters, driven by structural variations influenced by Mg2+ ions, challenging the assumption of unique folded states.

Contribution

The paper introduces a quantitative method using glass physics concepts to analyze smFRET data, demonstrating ergodicity breaking and conformational clustering in Holliday Junction dynamics.

Findings

Holliday Junction dynamics are effectively non-ergodic.

Conformational space is partitioned into kinetically disconnected clusters.

Mg2+ ions influence the structural heterogeneity and interconversion among clusters.

Abstract

A plausible consequence of rugged energy landscapes of biomolecules is that functionally competent folded states may not be unique, as is generally assumed. Indeed, molecule-to-molecule variations in the dynamics of enzymes and ribozymes under folding conditions have recently been identified in single molecule experiments. However, systematic quantification and the structural origin of the observed complex behavior remain elusive. Even for a relatively simple case of isomerization dynamics in Holliday Junctions (HJs), molecular heterogeneities persist over a long observation time (Tobs ~ 40 sec). Here, using concepts in glass physics and complementary clustering analysis, we provide a quantitative method to analyze the smFRET data probing the isomerization in HJ dynamics. We show that ergodicity of HJ dynamics is effectively broken; as a result, the conformational space of HJs is partitioned into a folding network of kinetically disconnected clusters. While isomerization dynamics in each cluster occurs rapidly as if the associated conformational space is fully sampled, distinct patterns of time series belonging to different clusters do not interconvert on Tobs. Theory suggests that persistent heterogeneity of HJ dynamics is a consequence of internal multiloops with varying sizes and flexibilities frozen by Mg2+ ions. An annealing experiment using Mg2+ pulse that changes the Mg2+ cocentration from high to low to high values lends support to this idea by explicitly showing that interconversions can be driven among trajectories with different patterns.