Dobson, J, Kumar, A orcid.org/0000-0001-8391-6812, Willis, LF orcid.org/0000-0001-6616-3716 et al. (8 more authors) (2017) Inducing protein aggregation by extensional flow. Proceedings of the National Academy of Sciences, 114 (18). pp. 4673-4678. ISSN 0027-8424
Abstract
Relative to other extrinsic factors, the effects of hydrodynamic flow fields on protein stability and conformation remain poorly understood. Flow-induced protein remodelling and/or aggregation is observed both in Nature and during the large-scale industrial manufacture of proteins. Despite its ubiquity, the relationships between the type and magnitude of hydrodynamic flow, a protein’s structure and stability and the resultant aggregation propensity are unclear. Here, we assess the effects of a defined and quantified flow-field dominated by extensional flow on the aggregation of bovine serum albumin (BSA), β2-microglobulin (β2m), granulocyte colony stimulating factor (G-CSF) and three monoclonal antibodies (mAbs). We show that the device induces protein aggregation after exposure to an extensional flow field for 0.36-1.8 ms, at concentrations as low as 0.5 mg mL-. In addition, we reveal that the extent of aggregation depends on the applied strain rate and the concentration, structural scaffold and sequence of the protein. Finally we demonstrate the in situ labelling of a buried cysteine residue in BSA during extensional stress. Together, these data indicate that an extensional flow readily unfolds thermodynamically and kinetically stable proteins, exposing previously sequestered sequences whose aggregation propensity determines the probability and extent of aggregation.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | This article is protected by copyright. All rights reserved. This is an author produced version of a paper published in the Proceedings of the National Academy of Sciences. Uploaded in accordance with the publisher's self-archiving policy. In order to comply with the publisher requirements the University does not require the author to sign a non-exclusive licence for this paper. |
Keywords: | extensional flow; aggregation; unfolding; bioprocessing; antibody |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Engineering & Physical Sciences (Leeds) > School of Mechanical Engineering (Leeds) > Institute of Engineering Thermofluids, Surfaces & Interfaces (iETSI) (Leeds) The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) > Biomolecular Mass Spectroscopy (Leeds) The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) > Structural Molecular Biology 2 (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 03 Apr 2017 16:35 |
Last Modified: | 13 Dec 2024 09:46 |
Published Version: | https://doi.org/10.1073/pnas.1702724114 |
Status: | Published |
Publisher: | National Academy of Sciences |
Identification Number: | 10.1073/pnas.1702724114 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:114410 |