Structural Evaluation of Protein/Metal Complexes via Native Electrospray Ultraviolet Photodissociation Mass Spectrometry
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Ultraviolet photodissociation (UVPD) has emerged as a promising tool to characterize proteins with regard to not only their primary sequences and post-translational modifications, but also their tertiary structures. In this study, three metal-binding proteins, Staphylococcal nuclease, azurin, and calmodulin, are used to demonstrate the use of UVPD to elucidate metal-binding regions via comparisons between the fragmentation patterns of apo (metal-free) and holo (metal-bound) proteins. The binding of staphylococcal nuclease to calcium was evaluated, in addition to a series of lanthanide(III) ions which are expected to bind in a similar manner as calcium. On the basis of comparative analysis of the UVPD spectra, the binding region for calcium and the lanthanide ions was determined to extend from residues 40-50, aligning with the known crystal structure. Similar analysis was performed for both azurin (interrogating copper and silver binding) and calmodulin (four calcium binding sites). This work demonstrates the utility of UVPD methods for determining and analyzing the metal binding sites of a variety of classes of proteins.
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Native MS provides information on protein complex subunit stoichiometry, metal binding, protein interaction partners, and lipid binding.
“Valuable information regarding protein complex subunit stoichiometry [42], metal-binding [43,44], protein interaction partners [45], and lipid binding [46] can be achieved using native MS.”
From Top-down proteomics: challenges, innovations, and applications in basic and clinical research (2020) · cites this paper as [43]
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Application of mass spectrometry techniques for analysis of higher order structure of proteins
Herath, D., Walls, K. N., Dollar, A. N., & Webb, I. (2026) · The Analyst · doi:10.1039/d6an00710d
DeFiglia, S. A., Kaiser, N. K., Blakney, G. T., Polák, M., & Håkansson, H. (2026) · Journal of the American Society for Mass Spectrometry · doi:10.1021/jasms.6c00260
- Authors
- Crittenden, C. M., Novelli, E. T., Mehaffey, M. R., Xu, G. N., Giles, D. H., Fies, W. A., Dalby, K. N., Webb, L. J., & Brodbelt, J. S.
- Affiliations
- The University of Texas at Austin (US)
- Texas Materials Institute (US)
- Corresponding author
- Brodbelt, Jennifer S.
- ORCID
- Venue
- Journal of the American Society for Mass Spectrometry
- Publisher
- American Chemical Society (ACS)
- Published
- 2020-04-10
- Volume
- 31
- Issue
- 5
- Pages
- 1140-1150
- ISSN
- 1044-0305 (ISSN-L)
- 1879-1123
- PMC
- PMC7386362
- OpenAlex
- W3015359929
- Cited by
- 24
- References
- 80
- Citations by year
- 2026: 3 · 2025: 1 · 2024: 4 · 2023: 4 · 2022: 4 · 2021: 5 · 2020: 3
- FWCI
- 1.53
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Licence
- Locations
- Journal of the American Society for Mass Spectrometry (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- Funders
- Division of Chemistry (CHE-1807215)
- Welch Foundation (F-1155, F-1390, F-1722, H-F-0001)
- Division of Molecular and Cellular Biosciences (MCB-1714555)
- National Institute of General Medical Sciences (R01 GM121714, R01 GM123252)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Keywords
- ultraviolet photodissociation, metal-binding proteins, metal binding sites, staphylococcal nuclease, azurin, calmodulin, calcium binding, lanthanide ions, copper binding, silver binding, protein tertiary structure, fragmentation patterns
- MeSH terms
- Amino Acid Sequence, Azurin, Binding Sites, Calcium, Calmodulin, Metals, Micrococcal Nuclease, Models, Molecular, Spectrometry, Mass, Electrospray Ionization, Lanthanoid Series Elements
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