Using Ion Mobility Spectrometry–Mass Spectrometry to Decipher the Conformational and Assembly Characteristics of the Hepatitis B Capsid Protein
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The structural and functional analysis of the core protein of hepatitis B virus is important for a full understanding of the viral life cycle and the development of novel therapeutic agents. The majority of the core protein (CP149) comprises the capsid assembly domain, and the C-terminal region (residues 150-183) is responsible for nucleic acid binding. Protein monomers associate to form dimeric structural subunits, and helices 3 and 4 (residues 50-111 of the assembly domain) have been shown to be important for this as they constitute the interdimer interface. Here, using mass spectrometry coupled with ion mobility spectrometry, we demonstrate the conformational flexibility of the CP149 dimer. Limited proteolysis was used to locate involvement in this feature to the C-terminal region. A genetically fused CP dimer was found to show decreased disorder, consistent with a more restricted C-terminus at the fusion junction. Incubation of CP149 dimer with heteroaryldihydropyrimidine-1, a small molecule known to interfere with the assembly process, was shown to result in oligomers different in shape to the capsid assembly-competent oligomers of the fused CP dimer. We suggest that heteroaryldihydropyrimidine-1 affects the dynamics of CP149 dimer in solution, likely affecting the ratio between assembly active and inactive states. Therefore, assembly of the less dynamic fused dimer is less readily misdirected by heteroaryldihydropyrimidine-1. These studies of the flexibility and oligomerization properties of hepatitis B virus core protein illustrate both the importance of C-terminal dynamics in function and the utility of gas-phase techniques for structural and dynamical biomolecular analysis.
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Native mass spectrometry combined with ion mobility is valuable for determining the size and shape of intermediate and mature capsid structures.
“When combined with ion mobility, a gas phase–based separation, native MS becomes a valuable technique for structural virologists to determine the size and shape of intermediate and mature capsid structures (Figure 2) (25–27).”
From The Impact of Mass Spectrometry–Based Proteomics on Fundamental Discoveries in Virology (2014) · cites this paper as [27]
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The Impact of Mass Spectrometry–Based Proteomics on Fundamental Discoveries in Virology
Greco, T. M., Diner, B. A., & Cristea, I. M. (2014) · Annual Review of Virology · doi:10.1146/annurev-virology-031413-085527 · Cited by 67 · see citing works
- Authors
- Shepherd, D. A., Holmes, K., Rowlands, D. J., Stonehouse, N. J., & Ashcroft, A. E.
- Affiliations
- University of Leeds (GB)
- Corresponding author
- Stonehouse, Nicola J.
- Ashcroft, Alison E.
- Venue
- Biophysical Journal
- Publisher
- Elsevier BV
- Published
- 2013-09
- Volume
- 105
- Issue
- 5
- Pages
- 1258-1267
- ISSN
- 0006-3495 (ISSN-L)
- PMC
- PMC3762345
- OpenAlex
- W2117756877
- Cited by
- 25
- References
- 44
- Citations by year
- 2025: 1 · 2022: 1 · 2021: 3 · 2020: 1 · 2019: 3 · 2018: 3 · 2017: 4 · 2016: 3 · 2015: 2 · 2014: 3 · 2013: 1
- FWCI
- 1.34
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- Locations
- Biophysical Journal (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- White Rose Research Online (University of Leeds, The University of Sheffield, University of York) (acceptedVersion)
- Funders
- Biotechnology and Biological Sciences Research Council (BB/E012558/1)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Health Sciences › Medicine › Epidemiology › Hepatitis B Virus Studies
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
- Keywords
- hepatitis B virus core protein, capsid assembly, protein flexibility, intrinsically disordered proteins, protein dimer, oligomerization, ion mobility mass spectrometry, limited proteolysis
- MeSH terms
- Amino Acid Sequence, Hepatitis B virus, Models, Molecular, Molecular Sequence Data, Pyridines, Pyrimidines, Protein Structure, Tertiary, Spectrometry, Mass, Electrospray Ionization, Capsid Proteins, Protein Multimerization
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