An in vivo platform to select and evolve aggregation-resistant proteins

Jessica S. Ebo, Janet C. Saunders, Paul W. A. Devine, Alice M. Gordon, Amy S. Warwick, Bob Schiffrin, Stacey E. Chin, Elizabeth England, James D. Button, Christopher Lloyd, Nicholas J. Bond, Alison E. Ashcroft, Sheena E. Radford, David C. Lowe () and David J. Brockwell ()
Additional contact information
Jessica S. Ebo: University of Leeds
Janet C. Saunders: University of Leeds
Paul W. A. Devine: University of Leeds
Alice M. Gordon: University of Leeds
Amy S. Warwick: University of Leeds
Bob Schiffrin: University of Leeds
Stacey E. Chin: AstraZeneca, Granta Park
Elizabeth England: AstraZeneca, Granta Park
James D. Button: AstraZeneca, Granta Park
Christopher Lloyd: AstraZeneca, Granta Park
Nicholas J. Bond: AstraZeneca, Granta Park
Alison E. Ashcroft: University of Leeds
Sheena E. Radford: University of Leeds
David C. Lowe: AstraZeneca, Granta Park
David J. Brockwell: University of Leeds

Nature Communications, 2020, vol. 11, issue 1, 1-12

Abstract: Abstract Protein biopharmaceuticals are highly successful, but their utility is compromised by their propensity to aggregate during manufacture and storage. As aggregation can be triggered by non-native states, whose population is not necessarily related to thermodynamic stability, prediction of poorly-behaving biologics is difficult, and searching for sequences with desired properties is labour-intensive and time-consuming. Here we show that an assay in the periplasm of E. coli linking aggregation directly to antibiotic resistance acts as a sensor for the innate (un-accelerated) aggregation of antibody fragments. Using this assay as a directed evolution screen, we demonstrate the generation of aggregation resistant scFv sequences when reformatted as IgGs. This powerful tool can thus screen and evolve ‘manufacturable’ biopharmaceuticals early in industrial development. By comparing the mutational profiles of three different immunoglobulin scaffolds, we show the applicability of this method to investigate protein aggregation mechanisms important to both industrial manufacture and amyloid disease.

Date: 2020
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DOI: 10.1038/s41467-020-15667-1

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