Immunoprecipitation and FRET-FLIM to Determine Metabolons on the Plant ER

Metabolons are protein complexes that contain all the enzymes necessary for a metabolic pathway but also scaffolding proteins. Such a structure allows efficient channeling of intermediate metabolites form one active site to the next and is highly advantageous for labile or toxic intermediates. Here we describe two methods currently used to identify metabolons via protein-protein interaction methodology: immunoprecipitations using GFP-Trap®_A beads to find novel interaction partners and potential metabolon components and FRET-FLIM to test for and quantify protein-protein interactions in planta.