Kissing loop interaction in adenine riboswitch: insights from umbrella sampling simulations

TL;DR

This study uses umbrella sampling simulations to investigate how ligand binding stabilizes the kissing loop interaction in an adenine riboswitch, providing detailed thermodynamic insights into its conformational regulation.

Contribution

It applies umbrella sampling with novel analysis methods to elucidate the ligand's role in stabilizing the kissing loop in an adenine riboswitch, advancing understanding of its structural mechanism.

Findings

Ligand stabilizes the kissing loop complex.

Simulation results align with experimental data.

Dependence on initialization protocol highlights need for improved sampling.

Abstract

Riboswitches are cis-acting regulatory RNA elements prevalently located in the leader sequences of bacterial mRNA. An adenine sensing riboswitch cis-regulates adeninosine deaminase gene (add) in Vibrio vulnificus. The structural mechanism regulating its conformational changes upon ligand binding mostly remains to be elucidated. In this open framework it has been suggested that the ligand stabilizes the interaction of the distal "kissing loop" complex. Using accurate full-atom molecular dynamics with explicit solvent in combination with enhanced sampling techniques and advanced analysis methods it could be possible to provide a more detailed perspective on the formation of these tertiary contacts. In this work, we used umbrella sampling simulations to study the thermodynamics of the kissing loop complex in the presence and in the absence of the cognate ligand. We enforced the breaking/formation of the loop-loop interaction restraining the distance between the two loops. We also assessed the convergence of the results by using two alternative initialization protocols. A structural analysis was performed using a novel approach to analyze base contacts. Our simulations qualitatively indicated that the ligand could stabilize the kissing loop complex. We also compared with previously published simulation studies. Kissing complex stabilization given by the ligand was compatible with available experimental data. However, the dependence of its value on the initialization protocol of the umbrella sampling simulations posed some questions on the quantitative interpretation of the results and called for better converged enhanced sampling simulations.