title: Measuring Hox-DNA binding by electrophoretic mobility shift analysis.
abstract: Understanding gene regulation by Hox transcription factors requires understanding the forces that underlie DNA binding by these proteins. Electrophoretic mobility shift analysis (EMSA) not only allows measurement of protein affinity and cooperativity but also permits visualization of differently migrating protein-DNA complexes, including complexes with different compositions or complexes with identical compositions yet assembled in different geometries. Furthermore, protein activity can be measured, allowing correction of binding constants for the percentage of protein that is properly folded and capable of binding DNA. Protocols for measuring protein activity and the equilibrium DNA-binding dissociation constant (K d) are provided. This versatile assay system can be adjusted based on specific needs to measure other parameters, including the kinetic association and dissociation constants (k a and k d) and the formation of heterologous protein-protein interactions.
authors.affiliation: Department of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, 77843-1114, USA.
authors.forename: Kelly
authors.initials: K
authors.lastname: Churion
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.forename: Hao-Ching
authors.initials: HC
authors.lastname: Hsiao
authors.forename: Kathleen S
authors.initials: KS
authors.lastname: Matthews
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Bondos
mesh_terms: D000818:Animals; D004247:DNA / Q000378:metabolism*; D004331:Drosophila melanogaster; D024202:Electrophoretic Mobility Shift Assay; D018398:Homeodomain Proteins / Q000378:metabolism*; D009841:Oligonucleotides / Q000378:metabolism; D011485:Protein Binding
chemical_list: D018398:Homeodomain Proteins; D009841:Oligonucleotides; D004247:DNA