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Chapters
- 00:05Title
- 01:30Transfection
- 02:20Cell Lysis
- 03:30Binding to Rabbit IgG-agarose
- 04:53TEV Protease Cleavage
- 05:48Binding to Immobilized Streptavidin Beads
- 07:50Biotin Elution of the Streptavidin Binging Peptide and Bait Protein
Tandem affinity purification is a robust approach for the identification of protein binding partners. As proof of concept, this methodology was applied to the well-characterized translation initiation factor eIF4E to co-precipitate the host cell factors involved in translation initiation. This method is easily adapted to any cellular or viral protein.
Tags
Protein Interacting PartnersTandem Affinity PurificationIdentification Of Interacting ProteinsYeast Two Hybrid SystemPull Down AssaysImmunoprecipitationMass Spectrometry IdentificationDouble-affinity Tag Mediated PurificationTAPEukaryotic Translation Initiation Factor EIF4ECellular Translation Factor EIF4EEIF4F ComplexCap-dependent Translation InitiationProtein G UnitsStreptavidin Binding PeptideTobacco Etch Virus (TEV) Protease Cleavage Sequence
