Polyelectrolyte interactions enable rapid association and dissociation in high-affinity disordered protein complexes

Madeleine B Borgia,Andrea Sottini,Benjamin Schuler,Birthe B Kragelund,Katrine Bugge,Daniel Nettels,Pétur O Heidarsson,Franziska Zosel,Alessandro Borgia,Robert B Best,Aritra Chowdhury

doi:10.1038/s41467-020-18859-x

Abstract

Highly charged intrinsically disordered proteins can form complexes with very high affinity in which both binding partners fully retain their disorder and dynamics, exemplified by the positively charged linker histone H1.0 and its chaperone, the negatively charged prothymosin α. Their interaction exhibits another surprising feature: The association/dissociation kinetics switch from slow two-state-like exchange at low protein concentrations to fast exchange at higher, physiologically relevant concentrations. Here we show that this change in mechanism can be explained by the formation of transient ternary complexes favored at high protein concentrations that accelerate the exchange between bound and unbound populations by orders of magnitude. Molecular simulations show how the extreme disorder in such polyelectrolyte complexes facilitates (i) diffusion-limited binding, (ii) transient ternary complex formation, and (iii) fast exchange of monomers by competitive substitution, which together enable rapid kinetics. Biological polyelectrolytes thus have the potential to keep regulatory networks highly responsive even for interactions with extremely high affinities.

Highlights

Charged intrinsically disordered proteins can form complexes with very high affinity in which both binding partners fully retain their disorder and dynamics, exemplified by the positively charged linker histone H1.0 and its chaperone, the negatively charged prothymosin α
To be able to probe the binding between ProTα and H1 from picomolar to micromolar concentrations, we used confocal single-molecule Förster resonance energy transfer (FRET) spectroscopy of freely diffusing molecules
By attaching a donor and an acceptor fluorophore to one of the two proteins, the binding of the unlabeled partner can be monitored by an increase in transfer efficiency, since the mutual charge screening of the oppositely charged intrinsically disordered proteins (IDPs) leads to chain compaction[12]

Summary

Introduction

Charged intrinsically disordered proteins can form complexes with very high affinity in which both binding partners fully retain their disorder and dynamics, exemplified by the positively charged linker histone H1.0 and its chaperone, the negatively charged prothymosin α. Their interaction exhibits another surprising feature: The association/dissociation kinetics switch from slow two-state-like exchange at low protein concentrations to fast exchange at higher, physiologically relevant concentrations. Molecular simulations show how the extreme disorder in such polyelectrolyte complexes facilitates (i) diffusion-limited binding, (ii) transient ternary complex formation, and (iii) fast exchange of monomers by competitive substitution, which together enable rapid kinetics. The underlying process of competitive substitution[14] via short-lived ternary complexes may be a means of keeping regulatory networks in the cell highly responsive even if individual binding partners have extreme affinities—a mechanism that could be widespread among IDP interactions

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Conclusion