Directed Assembly of Homopentameric Cholera Toxin B-Subunit Proteins into Higher-Order Structures Using Coiled-Coil Appendages

James F Ross, Gemma C Wildsmith, Michael Johnson, Daniel L Hurdiss, Kristian Hollingsworth, Rebecca F Thompson, Majid Mosayebi, Chi H Trinh, Emanuele Paci, Arwen R Pearson, Michael E Webb, W Bruce Turnbull

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

5 Citations (Scopus)
173 Downloads (Pure)

Abstract

The self-assembly of proteins into higher order structures is ubiquitous in living systems. It is also an essential process for the bottom-up creation of novel molecular architectures and devices for synthetic biology. However, the complexity of protein-protein interaction surfaces makes it challenging to mimic natural assembly processes in artificial systems. Indeed, many successful computationally designed protein assemblies are prescreened for "designability", limiting the choice of components. Here, we report a simple and pragmatic strategy to assemble chosen multisubunit proteins into more complex structures. A coiled-coil domain appended to one face of the pentameric cholera toxin B-subunit (CTB) enabled the ordered assembly of tubular supra-molecular complexes. Analysis of a tubular structure determined by X-ray crystallography has revealed a hierarchical assembly process that displays features reminiscent of the polymorphic assembly of polyomavirus proteins. The approach provides a simple and straightforward method to direct the assembly of protein building blocks which present either termini on a single face of an oligomer. This scaffolding approach can be used to generate bespoke supramolecular assemblies of functional proteins. Additionally, structural resolution of the scaffolded assemblies highlight "native-state" forced protein-protein interfaces, which may prove useful as starting conformations for future computational design.

Original languageEnglish
Pages (from-to)5211-5219
Number of pages9
JournalJournal of the American Chemical Society
Volume141
Issue number13
Early online date11 Mar 2019
DOIs
Publication statusPublished - 3 Apr 2019

Structured keywords

  • BrisSynBio
  • Bristol BioDesign Institute

Keywords

  • SYNTHETIC BIOLOGY

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