# Kinetic Traps in RNA Folding: Targeted Design of Frameshifting Element Mutants by Thermodynamic and Kinetic Analysis of the Chikungunya Virus

## TL;DR

This paper studies how the Chikungunya virus RNA folds during protein synthesis and shows how to design mutants that avoid kinetic traps to target specific RNA structures.

## Contribution

The paper introduces a computational framework for RNA design that accounts for both thermodynamic and kinetic factors to overcome kinetic traps.

## Key findings

- The FSE’s conformation is determined by competition between thermodynamic stability and cotranslational folding kinetics.
- The wildtype FSE is often trapped in simpler, less stable structures due to rapid formation during synthesis.
- An iterative design approach informed by kinetic analysis can drive the FSE into target conformations.

## Abstract

Chikungunya virus (CHIKV) employs a programmed ribosomal
frameshifting
element (FSE) to regulate the synthesis of its viral proteins, making
the FSE an attractive antiviral target. Yet the structural dynamics
that govern its function are complex and poorly understood, with multiple
folds discovered. Through computational analysis, we suggest that
the FSE’s conformation is determined by a competition between
thermodynamic stability and cotranslational folding kinetics. Using
an integrated computational pipeline, we map the FSE’s equilibrium
landscape, revealing a thermodynamically favored pseudoknot that emerges
only with sufficient flanking residues. We then use kinetic simulations
to show that, for the wildtype sequence, this pseudoknot is often
kinetically trapped in simpler, less stable stem loop structures that
form more rapidly during synthesis. Using this information, we rationally
design several mutants to target different folds in the FSE’s
repertoire. We demonstrate that while a purely thermodynamic design
can fail due to kinetic traps, an iterative design procedure, informed
by kinetic analysis, can drive the FSE onto a target conformation.
Our work explores conformational plasticity and multiple folding pathways
of the CHIKV FSE, shows how cotranslational kinetics influence the
fold-switching landscapes, establishes a computational framework for
kinetic-based RNA engineering, and highlights the importance of considering
folding pathways in the design of RNA-targeted therapeutics.