Affinity-purification coupled to mass spectrometry: basic principles and strategies
Abstract
Identifying the interactions established by a protein of interest can be a critical step in understanding its function. This is especially true when an unknown protein of interest is demonstrated to physically interact with proteins of known function. While many techniques have been developed to characterize protein-protein interactions, one strategy that has gained considerable momentum over the past decade for identification and quantification of protein-protein interactions, is affinity-purification followed by mass spectrometry (AP-MS). Here, we briefly review the basic principles used in affinity-purification coupled to mass spectrometry, with an emphasis on tools (both biochemical and computational), which enable the discovery and reporting of high quality protein-protein interactions.
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Limitations of the AP-MS experiment include the potential loss of transient or weakly bound complexes, the inherent noise associated with AP-MS relative to global proteomics analysis, and the short time point of the experiment, which precludes detection of degradation events with slow kinetics.
“Limitations of this experiment include the potential loss of transient or weakly bound complexes in the enrichment and subsequent wash steps, the inherent noise associated with AP-MS relative to global proteomics analysis (Dunham et al., 2012; Yugandhar et al., 2019) and the short time point of the experiment, which precludes detection of degradation events with slow kinetics.”
From Mapping the Degradable Kinome Provides a Resource for Expedited Degrader Development (2020) · cites this paper as Dunham et al., 2012
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- Authors
- Dunham, W. H., Mullin, M., & Gingras, A.
- Affiliations
- Mount Sinai Hospital (CA)
- Corresponding author
- Gingras, Anne‐Claude
- Venue
- PROTEOMICS
- Publisher
- Wiley
- Published
- 2012-05
- Volume
- 12
- Issue
- 10
- Pages
- 1576-1590
- ISSN
- 1615-9853 (ISSN-L)
- 1615-9861
- OpenAlex
- W1991953772
- Cited by
- 368
- References
- 144
- Citations by year
- 2026: 16 · 2025: 33 · 2024: 29 · 2023: 24 · 2022: 27 · 2021: 33 · 2020: 24 · 2019: 31 · 2018: 27 · 2017: 37 · 2016: 19 · 2015: 23 · 2014: 27 · 2013: 13 · 2012: 4
- FWCI
- 1.99
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- PROTEOMICS (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Canadian Institutes of Health Research (MOP 84314)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Bioinformatics and Genomic Networks
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Keywords
- affinity purification, protein-protein interaction, affinity purification mass spectrometry, protein-protein interaction network, protein complex identification
- MeSH terms
- Animals, Chromatography, Affinity, Humans, Proteins, Mass Spectrometry, Protein Interaction Mapping, Systems Biology
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