Molecular Simulations Matching Denaturation Experiments for N⁶-Methyladenosine

Abstract

Post-transcriptional modifications are crucial for RNA function and can affect its structure and dynamics. Force-field-based classical molecular dynamics simulations are a fundamental tool to characterize biomolecular dynamics, and their application to RNA is flourishing. Here, we show that the set of force-field parameters for N6-methyladenosine (m6A) developed for the commonly used AMBER force field does not reproduce duplex denaturation experiments and, specifically, cannot be used to describe both paired and unpaired states. Then, we use reweighting techniques to derive new parameters matching available experimental data. The resulting force field can be used to properly describe paired and unpaired m6A in both syn and anti conformation, which thus opens the way to the use of molecular simulations to investigate the effects of N6 methylations on RNA structural dynamics.

Alchemical Metadynamics: Adding Alchemical Variables to Metadynamics to Enhance Sampling in Free Energy Calculations

“…These profiles show that the syn conformation (central basin) is favored in the m 6 A nucleoside, whereas the anti conformation (lateral basins) is favored in the duplex. The final ΔΔ G values, which represent the amount by which the methylation disfavors the duplex, are consistent with those reported in ref within the respective statistical errors (Figure D).…”

“…(B) Residual free energy surface along the state index and the averaged torsional angle for the RNA duplex (C) Free energy computed as a function of η avg at fixed λ = 1, both for the RNA duplex (red) and for the m 6 A nucleoside (blue). (D) Comparison of ΔΔ G obtained with alchemical metadynamics (AM) and with Hamiltonian replica exchange (HREX) from Piomponi et al, with their respective statistical errors.…”

“…The barriers associated with the η angle are relevant to the kinetics of hybridization . In a previous paper, we studied this system using standard Hamiltonian replica exchange simulations and reparametrized partial charges, to fit thermodynamic data. , Specifically, we separately simulated the syn - and anti - conformations. We here show how to use alchemical metadynamics to recover the same information in a single simulation, and additionally obtain information about the isomerization barrier.…”

“…56 The barriers associated with the η angle are relevant to the kinetics of hybridization. 57 In a previous paper, 58 we studied this system using standard Hamiltonian replica exchange simulations and reparametrized partial charges, to fit thermodynamic data. 56,59 Specifically, we separately simulated the syn-and anti-conformations.…”

“…Preparation of Simulation Inputs. All the setups have been described extensively in Piomponi et al, 58 and are available on Zenodo (https://zenodo.org/record/6498021). Here, we consider the conversion of A to m 6 A in an isolated nucleoside (system A1 in ref 58) and in an RNA duplex (system A2 in ref 58, only the duplex system).…”