title: Synaptic MAGUK multimer formation is mediated by PDZ domains and promoted by ligand binding.
abstract: To examine the scaffolding properties of PSD-95, we have taken advantage of established ligand/PDZ domain interactions and developed a cell-based assay for investigating protein complex formation. This assay enables quantitative analysis of PDZ domain-mediated protein clustering using bimolecular fluorescence complementation (BiFC). Two nonfluorescent halves of EYFP were fused to C-terminal PDZ ligand sequences to generate probes that sense for PDZ domain binding grooves of adjacent (interacting) molecules. When these probes are brought into proximity by the PDZ domains of a multiprotein scaffold, a functional fluorescent EYFP molecule can be detected. We have used this system to examine the properties of selected PSD-95 variants and thereby delineated regions of importance for PSD-95 complex formation. Further analysis led to the finding that PSD-95 multimerization is PDZ domain-mediated and promoted by ligand binding.
authors: Neuroscience Research Center and Cluster of Excellence NeuroCure, Charité-Universitätsmedizin, 10117 Berlin, Germany.
authors: Nils
authors: N
authors: Rademacher
authors: Stella-Amrei
authors: SA
authors: Kunde
authors: Vera M
authors: VM
authors: Kalscheuer
authors: Sarah A
authors: SA
authors: Shoichet
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D005434:Flow Cytometry / Q000379:methods; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000032:analysis / Q000378:metabolism; D051528:Guanylate Kinases / Q000737:chemistry / Q000378:metabolism*; D008024:Ligands; D018613:Microscopy, Confocal / Q000379:methods; D008958:Models, Molecular; D054731:PDZ Domains; D011485:Protein Binding; D055503:Protein Multimerization; D013569:Synapses / Q000378:metabolism*
chemical_list: D005456:Fluorescent Dyes; D008024:Ligands; D051528:Guanylate Kinases