Cornwell, O, Ault, JR orcid.org/0000-0002-5131-438X, Bond, NJ et al. (2 more authors) (2021) An investigation of D76N β₂-microglobulin using protein footprinting and structural mass spectrometry. Journal of the American Society for Mass Spectrometry, 32 (7). pp. 1583-1592. ISSN 1044-0305
Abstract
NMR studies and X-ray crystallography have shown that the structures of the 99-residue amyloidogenic protein β₂-microglobulin (β₂m) and its more aggregation-prone variant, D76N, are indistinguishable, and hence, the reason for the striking difference in their aggregation propensities remains elusive. Here, we have employed two protein footprinting methods, hydrogen–deuterium exchange (HDX) and fast photochemical oxidation of proteins (FPOP), in conjunction with ion mobility-mass spectrometry, to probe the differences in conformational dynamics of the two proteins. Using HDX-MS, a clear difference in HDX protection is observed between these two proteins in the E-F loop (residues 70–77) which contains the D76N substitution, with a significantly higher deuterium uptake being observed in the variant protein. Conversely, following FPOP-MS only minimal differences in the level of oxidation between the two proteins are observed in the E–F loop region, suggesting only modest side-chain movements in that area. Together the HDX-MS and FPOP-MS data suggest that a tangible perturbation to the hydrogen-bonding network in the E–F loop has taken place in the D76N variant and furthermore illustrate the benefit of using multiple complementary footprinting methods to address subtle, but possibly biologically important, differences between highly similar proteins.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2021 American Society for Mass Spectrometry. Published by American Chemical Society. All rights reserved. This publication is licensed under CC-BY 4.0. |
Keywords: | β₂m; HDX; FPOP; amyloid; D76N; protein conformation; structural mass spectrometry |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | 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 (Leeds) |
Funding Information: | Funder Grant number BBSRC (Biotechnology & Biological Sciences Research Council) BB/K000659/1 BBSRC (Biotechnology & Biological Sciences Research Council) BB/E012558/1 EU - European Union 322408 |
Depositing User: | Symplectic Publications |
Date Deposited: | 15 Feb 2021 14:10 |
Last Modified: | 22 Mar 2024 10:22 |
Status: | Published |
Publisher: | American Chemical Society |
Identification Number: | 10.1021/jasms.0c00438 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:170895 |