ArticleNature protocols2014
Ecto-Fc MS identifies ligand-receptor interactions through extracellular domain Fc fusion protein baits and shotgun proteomic analysis.
Article in Nature protocols, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 15 papers.
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Who cites it
15 citing papers in PubMed, 28 citations in OpenAlex.
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- Synaptogenic activity of the axon guidance molecule Robo2 underlies hippocampal circuit function.Cell reports · 2021Article
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- Synapse type-specific proteomic dissection identifies IgSF8 as a hippocampal CA3 microcircuit organizer.Nature communications · 2020Article
- Secreted amyloid-β precursor protein functions as a GABAScience (New York, N.Y.) · 2019Article
- An Input-Specific Orphan Receptor GPR158-HSPG Interaction Organizes Hippocampal Mossy Fiber-CA3 Synapses.Neuron · 2018Article
- EpCAM ectodomain EpEX is a ligand of EGFR that counteracts EGF-mediated epithelial-mesenchymal transition through modulation of phospho-ERK1/2 in head and neck cancers.PLoS biology · 2018Article
- Subnanogram proteomics: impact of LC column selection, MS instrumentation and data analysis strategy on proteome coverage for trace samples.International journal of mass spectrometry · 2018Article
- Structural Mechanism for Modulation of Synaptic Neuroligin-Neurexin Signaling by MDGA Proteins.Neuron · 2017Article
- Proteomic Analysis of Protein Turnover by Metabolic Whole Rodent Pulse-Chase Isotopic Labeling and Shotgun Mass Spectrometry Analysis.Methods in molecular biology (Clifton, N.J.) · 2016Article
- Specification of synaptic connectivity by cell surface interactions.Nature reviews. Neuroscience · 2016Review
- Article
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Authors and funding
6 authors at 5 institutions in 3 countries.
Funding
Abstract
Ligand-receptor interactions represent essential biological triggers that regulate many diverse and important cellular processes. We have developed a discovery-based proteomic biochemical protocol that couples affinity purification with multidimensional liquid chromatographic tandem mass spectrometry (LCLC-MS/MS) and bioinformatic analysis. Compared with previous approaches, our analysis increases sensitivity, shortens analysis duration and boosts comprehensiveness. In this protocol, receptor extracellular domains are fused with the Fc region of IgG to generate fusion proteins that are purified from transfected HEK293T cells. These 'ecto-Fcs' are coupled to protein A beads and serve as baits for binding assays with prey proteins extracted from rodent brain. After capture, the affinity-purified proteins are digested into peptides and comprehensively analyzed by LCLC-MS/MS with ion-trap mass spectrometers. In 4 working days, this protocol can generate shortlists of candidate ligand-receptor protein-protein interactions. Our 'ecto-Fc MS' approach outperforms antibody-based approaches and provides a reproducible and robust framework for identifying extracellular ligand-receptor interactions.
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Registered trials
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.