Vanhinsbergh, C.J., Criscuolo, A., Sutton, J.N. et al. (4 more authors) (2022) Characterization and sequence mapping of large RNA and mRNA therapeutics using mass spectrometry. Analytical Chemistry, 94 (20). pp. 7339-7349. ISSN 0003-2700
Abstract
Large RNA including mRNA (mRNA) has emerged as an important new class of therapeutics. Recently, this has been demonstrated by two highly efficacious vaccines based on mRNA sequences encoding for a modified version of the SARS-CoV-2 spike protein. There is currently significant demand for the development of new and improved analytical methods for the characterization of large RNA including mRNA therapeutics. In this study, we have developed an automated, high-throughput workflow for the rapid characterization and direct sequence mapping of large RNA and mRNA therapeutics. Partial RNase digestions using RNase T1 immobilized on magnetic particles were performed in conjunction with high-resolution liquid chromatography–mass spectrometry analysis. Sequence mapping was performed using automated oligoribonucleotide annotation and identifications based on MS/MS spectra. Using this approach, a >80% sequence of coverage of a range of large RNAs and mRNA therapeutics including the SARS-CoV-2 spike protein was obtained in a single analysis. The analytical workflow, including automated sample preparation, can be completed within 90 min. The ability to rapidly identify, characterize, and sequence map large mRNA therapeutics with high sequence coverage provides important information for identity testing, sequence validation, and impurity analysis.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2022 The Authors. Published by American Chemical Society. This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (http://creativecommons.org/licenses/by/4.0) |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Engineering (Sheffield) > Department of Chemical and Biological Engineering (Sheffield) |
Funding Information: | Funder Grant number BIOTECHNOLOGY AND BIOLOGICAL SCIENCES RESEARCH COUNCIL BB/M012166/1 |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 20 May 2022 12:34 |
Last Modified: | 02 Feb 2023 14:38 |
Status: | Published |
Publisher: | American Chemical Society |
Refereed: | Yes |
Identification Number: | 10.1021/acs.analchem.2c00765 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:187112 |