# Consequences of mRNA Secondary Structure on Stability against both Hydrolysis and Aggregation: The Role of Electrostatic, π–π Stacking, and Thermal Effects

## TL;DR

This paper explores how mRNA's secondary structure affects its stability against breakdown and clumping, revealing new insights into the role of electrostatic and stacking forces.

## Contribution

The study links mRNA aggregation and autohydrolysis through secondary structure effects, introducing a new model involving electrostatic and π–π stacking interactions.

## Key findings

- mRNA secondary structure stabilizes against autohydrolysis and aggregation.
- Guanidine-HCl and temperature destabilize mRNA structure, increasing aggregation and hydrolysis.
- A new model explains aggregation via electrostatic and π–π stacking effects.

## Abstract

The seemingly unrelated massive aggregation of free mRNA
under
certain solution conditions and the well-known autohydrolysis of mRNA
are actually both closely linked through its secondary and possibly
tertiary structure (s/t). This hypothesis posits that s/t partially
stabilizes mRNA against both autohydrolysis and massive aggregation.
Destabilization of s/t via denaturant guanidine-HCl (Gd), or temperature,
has profound effects on both aggregation rates and final degree of
autohydrolysis. These denaturant effects occurred for a variety of
mRNA, ranging from 700 to 3000 nucleotides but showed very different
quantitative behavior among themselves, suggesting some of the methods
presented here might help characterize mRNA stability and robustness.
Light scattering monitoring during dialysis of mRNA against Gd revealed
an “aggregation window”, over 0.5–3 M Gd, whereas
dialyzing against a nondenaturing electrolyte (NaCl) showed semireversible
monotonic increase of aggregation up to 4 M. Massive aggregation of
mRNA in solutions with monovalent ions and in denaturing solutions
has not been previously reported. A phenomenological model involving
intermolecular electrostatic repulsion and attractions due chiefly
to π–π stacking helps interpret the various phenomena.