Cited by 24Mass Spectrometry Techniques and ApplicationsProtein Structure and DynamicsAdvanced Proteomics Techniques and Applications

Structural Evaluation of Protein/Metal Complexes via Native Electrospray Ultraviolet Photodissociation Mass Spectrometry

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Crittenden, Christopher M., Novelli, Elisa T., Mehaffey, M. Rachel, Xu, Gulan N., Giles, David H., Fies, Whitney A., Dalby, Kevin N., Webb, Lauren J. +1 more·2020·Journal of the American Society for Mass Spectrometry

Abstract

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.”

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  1. [57]Ferapontova EE; Shleev S; Ruzgas T; Stoica L; Christenson A; Tkac J; Yaropolov AI; Gorton L Direct Electrochemistry of Proteins and Enzymes In Paleček E; Scheller F; Wang J, Eds. Perspectives in Bioanalysis; pp 517– 598; Elsevier, 2005.

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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.
Venue
Journal of the American Society for Mass Spectrometry
Published
2020-04-10
Volume
31
Issue
5
Pages
1140-1150
ISSN
  • 1044-0305 (ISSN-L)
  • 1879-1123
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
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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