Analyzing protein complexes in Drosophila with tandem affinity purification–mass spectrometry
- 9 February 2005
- journal article
- research article
- Published by Wiley in Developmental Dynamics
- Vol. 232 (3), 827-834
- https://doi.org/10.1002/dvdy.20272
Abstract
We describe the application of tandem affinity purification–mass spectrometry (TAP‐MS) to the study of protein complexes in Drosophila. We have constructed vectors for inducible expression of TAP‐tagged fusion proteins in Drosophila cultured cells and in vivo. Using these vectors, we tagged, as a paradigm, several components of the Notch signaling pathway, isolated protein complexes containing the baits and associated proteins from cells and embryos, and identified the subunits by liquid chromatography–tandem mass spectrometry (LC‐MS/MS). Several known interactions involving Notch pathway elements were confirmed, and many novel potential interactions were uncovered. For some of the novel associations, we validated the interaction genetically and biochemically. We conclude that TAP, in combination with MS, can be used as an effective method for the studies of the Drosophila proteome. Developmental Dynamics 232:827–834, 2005.Keywords
This publication has 27 references indexed in Scilit:
- The Drosophila F Box Protein Archipelago Regulates dMyc Protein Levels In VivoCurrent Biology, 2004
- A physical and functional map of the human TNF-α/NF-κB signal transduction pathwayNature, 2004
- Mastermind mediates chromatin-specific transcription and turnover of the Notch enhancer complexGenes & Development, 2002
- Functional organization of the yeast proteome by systematic analysis of protein complexesNature, 2002
- The Tandem Affinity Purification (TAP) Method: A General Procedure of Protein Complex PurificationMethods, 2001
- Uncoupling cadherin-based adhesion from wingless signalling in DrosophilaNature, 1996
- An activated Notch receptor blocks cell-fate commitment in the developing Drosophila eyeNature, 1993
- Specific EGF repeats of Notch mediate interactions with Delta and serrate: Implications for notch as a multifunctional receptorCell, 1991
- Molecular interactions between the protein products of the neurogenic loci Notch and Delta, two EGF-homologous genes in DrosophilaCell, 1990
- Transposition of Cloned P Elements into Drosophila Germ Line ChromosomesScience, 1982