The RNA chaperone StpA enables fast RNA refolding by destabilization of mutually exclusive base pairs within competing secondary structure elements.

Katharina F Hohmann,Alexander Heckel,Boris Fürtig,Anja Blümler

doi:10.1093/nar/gkab876

Abstract

In bacteria RNA gene regulatory elements refold dependent on environmental clues between two or more long-lived conformational states each associated with a distinct regulatory state. The refolding kinetics are strongly temperature-dependent and especially at lower temperatures they reach timescales that are biologically not accessible. To overcome this problem, RNA chaperones have evolved. However, the precise molecular mechanism of how these proteins accelerate RNA refolding reactions remains enigmatic. Here we show how the RNA chaperone StpA of Escherichia coli leads to an acceleration of a bistable RNA’s refolding kinetics through the selective destabilization of key base pairing interactions. We find in laser assisted real-time NMR experiments on photocaged bistable RNAs that the RNA chaperone leads to a two-fold increase in refolding rates at low temperatures due to reduced stability of ground state conformations. Further, we can show that upon interaction with StpA, base pairing interactions in the bistable RNA are modulated to favor refolding through the dominant pseudoknotted transition pathway. Our results shed light on the molecular mechanism of the interaction between RNA chaperones and bistable RNAs and are the first step into a functional classification of chaperones dependent on their biophysical mode of operation.

Highlights

Modulation of gene expression in bacteria can be provided by RNA regulators
In order to determine the influence of the RNA Chaperone StpA-C-terminal domain (CTD) on the thermodynamics of the bistable RNA, we investigated the conformational equilibrium of the RNA by NMR both in absence and presence of different amounts of the protein
We can show that the RNA chaperone StpA-CTD, that is known to bind weakly and transiently to RNA [28], binds with a KD in the lower decadic ␮M range and promotes the refolding reaction by modulation of the underlying thermodynamics of the bistable system

Summary

Introduction

Modulation of gene expression in bacteria can be provided by RNA regulators. These RNAs functionally exploit their inherent structural plasticity that is founded in their natural ability to adopt more than a single stable conformation.Thereby they constitute a bi- or even multi-stable system. Modulation of gene expression in bacteria can be provided by RNA regulators These RNAs functionally exploit their inherent structural plasticity that is founded in their natural ability to adopt more than a single stable conformation. Within the regulation of gene expression, the conformational states of a bistable system are typically linked to two functional states (ON and OFF) [1,2] Prime examples for such biological bistable RNAs are RNA thermometers and riboswitches found in the 5 - or 3 -untranslated regions of mRNAs [2,3,4,5,6]. The involved states have similar thermodynamic stability rendering the difference in free energy close to zero, and thereby facilitating a change in the population ratio by environmental clues

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Results

Conclusion