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An in vivo platform to select and evolve aggregation-resistant proteins
Jessica S Ebo1,2, Janet C Saunders1,2,3,3, Paul W A Devine1,2,3
1Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, LS2 9JT, UK.
Nature Communications
|April 15, 2020
Summary
Scientists developed a new assay to detect and prevent protein aggregation in biopharmaceuticals. This method aids in creating more stable antibody fragments, improving drug manufacturing and potentially treating diseases like amyloidosis.
Area of Science:
- Biochemistry
- Biotechnology
- Protein Engineering
Background:
- Protein biopharmaceuticals are crucial but prone to aggregation during production and storage.
- Predicting aggregation is challenging as it's linked to non-native states, not just thermodynamic stability.
- Current methods for identifying stable protein sequences are inefficient.
Purpose of the Study:
- To develop a novel screening method for identifying and engineering aggregation-resistant protein sequences.
- To create a tool for early-stage industrial development of 'manufacturable' biopharmaceuticals.
- To investigate protein aggregation mechanisms relevant to biopharmaceutical manufacturing and amyloid diseases.
Main Methods:
- An assay was engineered in E. coli periplasm linking protein aggregation to antibiotic resistance.
- This assay functions as a direct sensor for the innate aggregation of antibody fragments.
- Directed evolution was employed using this assay to screen for aggregation-resistant single-chain variable fragments (scFvs).
Main Results:
- The assay successfully identified and facilitated the evolution of aggregation-resistant scFv sequences.
- Engineered aggregation-resistant sequences were confirmed when reformatted as immunoglobulin Gs (IgGs).
- Comparative analysis of mutational profiles across different immunoglobulin scaffolds provided insights into aggregation mechanisms.
Conclusions:
- The developed assay is a powerful tool for screening and evolving aggregation-resistant biopharmaceuticals.
- This method enables early-stage identification of 'manufacturable' protein therapeutics.
- The approach is applicable to understanding protein aggregation in both industrial contexts and diseases like amyloidosis.
