ArticleMolecular & cellular proteomics : MCP2015
Accurate protein complex retrieval by affinity enrichment mass spectrometry (AE-MS) rather than affinity purification mass spectrometry (AP-MS).
Article in Molecular & cellular proteomics : MCP, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 132 papers.
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132 citing papers in PubMed, 272 citations in OpenAlex.
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- There and back again: a cell biologist's journey from organelles to molecules.Biological chemistry · 2025Review
- Analysis of the Porcine Reproductive and Respiratory Syndrome Virus Nucleocapsid Interactome.Journal of proteome research · 2025Article
- Systematic metaproteomics mapping reveals functional and ecological landscapes of Ex vivo human gut microbiota responses to therapeutic drugs.Nature communications · 2025Article
- The Developmental Environment Mediates Adult Seminal Proteome Allocation in Male Drosophila melanogaster.Molecular ecology · 2025Article
- TDP-43 pathology is associated with divergent protein profiles in ALS brain and spinal cord.Acta neuropathologica communications · 2025Article
- Comparative Analysis of Protein Extraction Protocols for Olive Leaf Proteomics: Insights into Differential Protein Abundance and Isoelectric Point Distribution.ACS agricultural science & technology · 2025Article
- Affinity Purification Mass Spectrometry on the Orbitrap-Astral Mass Spectrometer Enables High-Throughput Protein-Protein Interaction Mapping.Journal of proteome research · 2025Article
- Article
- Exploring the anticancer mechanism of cardiac glycosides using proteome integral solubility alteration approach.Cancer medicine · 2024Article
- Inheritance of H3K9 methylation regulates genome architecture in Drosophila early embryos.The EMBO journal · 2024Article
- Quantitative proteomics reveals CLR interactome in primary human cells.The Journal of biological chemistry · 2024Article
- Hepatitis B virus RNAs co-opt ELAVL1 for stabilization and CRM1-dependent nuclear export.PLoS pathogens · 2024Article
72 more citing papers are in PubMed but not listed here.
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Protein-protein interactions are fundamental to the understanding of biological processes. Affinity purification coupled to mass spectrometry (AP-MS) is one of the most promising methods for their investigation. Previously, complexes were purified as much as possible, frequently followed by identification of individual gel bands. However, todays mass spectrometers are highly sensitive, and powerful quantitative proteomics strategies are available to distinguish true interactors from background binders. Here we describe a high performance affinity enrichment-mass spectrometry method for investigating protein-protein interactions, in which no attempt at purifying complexes to homogeneity is made. Instead, we developed analysis methods that take advantage of specific enrichment of interactors in the context of a large amount of unspecific background binders. We perform single-step affinity enrichment of endogenously expressed GFP-tagged proteins and their interactors in budding yeast, followed by single-run, intensity-based label-free quantitative LC-MS/MS analysis. Each pull-down contains around 2000 background binders, which are reinterpreted from troubling contaminants to crucial elements in a novel data analysis strategy. First the background serves for accurate normalization. Second, interacting proteins are not identified by comparison to a single untagged control strain, but instead to the other tagged strains. Third, potential interactors are further validated by their intensity profiles across all samples. We demonstrate the power of our AE-MS method using several well-known and challenging yeast complexes of various abundances. AE-MS is not only highly efficient and robust, but also cost effective, broadly applicable, and can be performed in any laboratory with access to high-resolution mass spectrometers.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.