Trajectory Taken by Dimeric Cu/Zn Superoxide Dismutase through the Protein Unfolding and Dissociation Landscape Is Modulated by Salt Bridge Formation
| Autor: | Celine Kelso, J. Andrew Aquilina, Justin J. Yerbury, Luke McAlary, S. P. Fitzgerald, Justin L. P. Benesch, Julian A. Harrison |
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| Rok vydání: | 2019 |
| Předmět: |
Models
Molecular 010402 general chemistry Mass spectrometry 01 natural sciences Mass Spectrometry Dissociation (chemistry) Analytical Chemistry Ion chemistry.chemical_compound Superoxide Dismutase-1 Humans Point Mutation Folding funnel Protein Unfolding Tandem 010401 analytical chemistry Energy landscape Recombinant Proteins 0104 chemical sciences Kinetics Crystallography Monomer chemistry Thermodynamics Ampicillin Dimerization Macromolecule |
| Zdroj: | Analytical Chemistry. 92:1702-1711 |
| ISSN: | 1520-6882 0003-2700 |
| DOI: | 10.1021/acs.analchem.9b01699 |
| Popis: | Native mass spectrometry (MS) is a powerful means for studying macromolecular protein assemblies, including accessing activated states. However, much remains to be understood about what governs which regions of the protein (un)folding funnel, which can be explored by activation of protein ions in a vacuum. Here, we examine the trajectory that Cu/Zn superoxide dismutase (SOD1) dimers take over the unfolding and dissociation free energy landscape in a vacuum. We examined wild-type SOD1 and six disease-related point mutants by using tandem MS and ion-mobility MS as a function of collisional activation. For six of the seven SOD1 variants, increasing activation prompted dimers to transition through two unfolding events and dissociate symmetrically into monomers with (as near as possible) equal charges. The exception was G37R, which proceeded only through the first unfolding transition and displayed a much higher abundance of asymmetric products. Supported by the observation that ejected asymmetric G37R monomers were more compact than symmetric G37R ones, we localized this effect to the formation of a gas-phase salt bridge in the first activated conformation. To examine the data quantitatively, we applied Arrhenius-type analysis to estimate the barriers on the corresponding free energy landscape. This reveals a heightening of the barrier to unfolding in G37R by >5 kJ/mol-1 over the other variants, consistent with expectations for the strength of a salt bridge. Our work demonstrates weaknesses in the simple general framework for understanding protein complex dissociation in a vacuum and highlights the importance of individual residues, their local environment, and specific interactions in governing product formation. |
| Databáze: | OpenAIRE |
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