title: Identification of DNA-protein complexes using an improved, combined western blotting-electrophoretic mobility shift assay (WEMSA) with a fluorescence imaging system.
abstract: Electrophoretic Mobility Shift Assays (EMSAs) are used to detect DNA-protein interactions. With this type of assay it is difficult to distinguish between specific and non-specific DNA-protein complexes or to define which protein binds to the DNA. Here we describe a novel Western blot-combined EMSA (WEMSA) variant for a fluorescence imaging system which permits easy identification of specific DNA-protein-complexes. This method also allows investigation of several DNA-protein complexes in parallel. We have identified and distinguished clearly between the SP1- and EGR1-DNA-protein complexes which exhibit overlapping binding to the GC-BOX of the mPGES-1 promoter.
authors.affiliation: Pharmazentrum frankfurt/ZAFES, Institute of Clinical Pharmacology, Goethe-University Frankfurt, Theodor Stern Kai 7, 60590 Frankfurt/Main, Germany. Klaus-Deckmann@gmx.de
authors.forename: Klaus
authors.initials: K
authors.lastname: Deckmann
authors.forename: Florian
authors.initials: F
authors.lastname: Rörsch
authors.forename: Gerd
authors.initials: G
authors.lastname: Geisslinger
authors.forename: Sabine
authors.initials: S
authors.lastname: Grösch
mesh_terms: D001483:Base Sequence; D015153:Blotting, Western / Q000379:methods*; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D051766:Early Growth Response Protein 1 / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods*; D005453:Fluorescence; D006367:HeLa Cells; D006801:Humans; D019746:Intramolecular Oxidoreductases / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D000071876:Prostaglandin-E Synthases; D011485:Protein Binding; D016329:Sp1 Transcription Factor / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D016329:Sp1 Transcription Factor; D004247:DNA; D019746:Intramolecular Oxidoreductases; C000605862:PTGES protein, human; D000071876:Prostaglandin-E Synthases