Combining native and ‘omics’ mass spectrometry to identify endogenous ligands bound to membrane proteins
Open accessAbstract
Ligands bound to protein assemblies provide critical information for function, yet are often difficult to capture and define. Here we develop a top-down method, 'nativeomics', unifying 'omics' (lipidomics, proteomics, metabolomics) analysis with native mass spectrometry to identify ligands bound to membrane protein assemblies. By maintaining the link between proteins and ligands, we define the lipidome/metabolome in contact with membrane porins and a mitochondrial translocator to discover potential regulators of protein function.
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Direct infusion native MS workflows can identify metabolites bound to proteins by MS/MS.
“While direct infusion native MS workflows have been developed that can identify metabolites bound to proteins by MS/MS29”
From Native metabolomics identifies the rivulariapeptolide family of protease inhibitors (2022) · cites this paper as [29]
MS4 analysis can identify both the proteoform state and the associated ligand structure.
“For example, Gault et al. demonstrated the use of MS4 analysis to identify both the proteoform state and the associated ligand structure [123].”
From Top-down proteomics: challenges, innovations, and applications in basic and clinical research (2020) · cites this paper as [123]
MS/MS analysis has been used to study protein complexes and protein-ligand binding.
“Beyond primary sequence coverage, MS/MS analysis has been utilized to study protein complexes [122] and protein-ligand binding [123].”
From Top-down proteomics: challenges, innovations, and applications in basic and clinical research (2020) · cites this paper as [123]
With the Nativeomics approach, endogenous ligands such as lipids and drug binding could be identified for several membrane proteins.
“Using the ‘Nativeomics’ approach, endogenous ligands such as lipids and even drug binding could be identified as demonstrated for several membrane proteins.”
From Top-down proteomics: challenges, innovations, and applications in basic and clinical research (2020) · cites this paper as [123]
The first step of complex-down experiments is to transfer a noncovalent complex from the solution phase to the gas phase without excessive disruption of its higher-order structure.
“The first step of these experiments is to transfer a noncovalent complex from the solution phase to the gas phase without excessive disruption of its higher-order structure.103”
From Higher-order structural characterisation of native proteins and complexes by top-down mass spectrometry (2020) · cites this paper as [103]
Efficient sequencing can be combined with obtaining information on noncovalent complex stoichiometry through MSn (n = 3 or more) analysis of complexes ionised under non-denaturing conditions.
“Efficient sequencing can be combined with obtaining information on noncovalent complex stoichiometry through MSn (n = 3 or more) analysis of complexes ionised under non-denaturing conditions.”
From Higher-order structural characterisation of native proteins and complexes by top-down mass spectrometry (2020) · cites this paper as [103]
Robinson and co-workers applied the complex-down workflow to show that MSn analysis was able to identify a lipid molecule bound to the outer mitochondrial membrane translocator protein complex.
“Recently, Robinson and co-workers applied this workflow to show that MSn analysis was able to identify a lipid molecule that was bound to the outer mitochondrial membrane translocator protein complex.103”
From Higher-order structural characterisation of native proteins and complexes by top-down mass spectrometry (2020) · cites this paper as [103]
The lipid bound to the outer mitochondrial membrane translocator protein complex was phosphatidylethanolamine 34:1, which fit well with the existing crystal structure.
“The lipid was found to be a phosphatidylethanolamine 34:1, which fit well with the existing crystal structure.”
From Higher-order structural characterisation of native proteins and complexes by top-down mass spectrometry (2020) · cites this paper as [103]
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- Authors
- Gault, J., Liko, I., Landreh, M., Shutin, D., Bolla, J. R., Jefferies, D., Agasid, M., Yen, H.-Y., Ladds, M. J. G. W., Lane, D. P., Khalid, S., Mullen, C., Remes, P. M., Huguet, R., McAlister, G., Goodwin, M., Viner, R., Syka, J. E. P., & Robinson, C. V.
- Affiliations
- University of Oxford (GB)
- Karolinska Institutet (SE)
- University of Southampton (GB)
- Thermo Fisher Scientific (United States) (US)
- Corresponding author
- Gault, Joseph
- Robinson, Carol V.
- ORCID
- Venue
- Nature Methods
- Published
- 2020-05
- Volume
- 17
- Issue
- 5
- Pages
- 505-508
- ISSN
- 1548-7091 (ISSN-L)
- 1548-7105
- PMC
- PMC7332344
- OpenAlex
- W3021994240
- Cited by
- 171
- References
- 47
- Citations by year
- 2026: 10 · 2025: 26 · 2024: 26 · 2023: 36 · 2022: 34 · 2021: 26 · 2020: 12
- FWCI
- 10.31
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Nature Methods (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- ePrints Soton (University of Southampton) (submittedVersion)
- Funders
- Case Western Reserve University
- University of Canterbury
- McGill University
- Wellcome Trust (104633, 104633/Z/14/Z)
- McGill University Health Centre
- University of Southampton
- University of Oxford
- Stiftelsen för Strategisk Forskning
- Medical Research Council (MR/N020413/1)
- Engineering and Physical Sciences Research Council (EP/R029407/1, R029407, EP/P511377/1)
- H2020 European Research Council (641317)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolomics and Mass Spectrometry Studies
- Keywords
- native mass spectrometry, membrane protein assembly, endogenous ligands, protein-ligand complex, lipidomics, proteomics, metabolomics, mitochondrial translocase, top-down mass spectrometry, ligand identification
- MeSH terms
- Humans, Ligands, Lipids, Membrane Proteins, Mass Spectrometry, Proteome, Metabolome
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