Maturation and Conformational Switching of a De Novo Designed Phase-Separating Polypeptide

Alex Hilditch*, Andrey Romanyuk, Lorna R Hodgson, Judith M Mantell, Chris R Neal, Paul Verkade, Richard J Obexer, Louise C. Serpell, Jennifer McManus*, Dek N Woolfson*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

Abstract

Cellular compartments formed by biomolecular condensation are widespread features of cell biology. These organelle-like assemblies compartmentalize macromolecules dynamically within the crowded intracellular environment. However, the intermolecular interactions that produce condensed droplets may also create arrested states and potentially pathological assemblies such as fibers, aggregates, and gels through droplet maturation. Protein liquid–liquid phase separation is a metastable process, so maturation may be an intrinsic property of phase-separating proteins, where nucleation of different phases or states arises in supersaturated condensates. Here, we describe the formation of both phase-separated droplets and proteinaceous fibers driven by a de novo designed polypeptide. We characterize the formation of supramolecular fibers in vitro and in bacterial cells. We show that client proteins can be targeted to the fibers in cells using a droplet-forming construct. Finally, we explore the interplay between phase separation and fiber formation of the de novo polypeptide, showing that the droplets mature with a post-translational switch to largely β conformations, analogous to models of pathological phase separation.
Original languageEnglish
Pages (from-to)10240–10245
Number of pages6
JournalJournal of the American Chemical Society
Volume146
Issue number15
Early online date5 Apr 2024
DOIs
Publication statusPublished - 17 Apr 2024

Bibliographical note

Publisher Copyright:
© 2024 The Authors. Published by American Chemical Society.

Structured keywords

  • Max Planck Bristol
  • Bristol BioDesign Institute

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