title: A fluorescence polarization assay for identifying ligands that bind to vascular endothelial growth factor.
abstract: Vascular endothelial growth factor (VEGF) is a homodimeric proangiogenic protein that induces endothelial cell migration and proliferation primarily through interactions with its major receptors, VEGFR-1 and VEGFR-2. Inhibitors of one or both of these VEGF-receptor interactions could be beneficial as therapeutics for diseases caused by dysfunctional angiogenesis (e.g., cancer). Others have reported small peptides that bind to the VEGF dimer at surface regions that are recognized by the receptors. Here we report the development of a fluorescence polarization assay based on the binding to VEGF of a derivative of one of these peptides that has been labeled with BODIPY-tetramethylrhodamine (BODIPY(TMR)). This 384-well format assay is tolerant to dimethyl sulfoxide (DMSO, up to 4% [v/v]) and has a Z' factor of 0.76, making it useful for identifying molecules that associate with the receptor-binding surface of the VEGF dimer.
authors.affiliation: Department of Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Kimberly J
authors.initials: KJ
authors.lastname: Peterson
authors.forename: Jack D
authors.initials: JD
authors.lastname: Sadowsky
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Scheef
authors.forename: Soumen
authors.initials: S
authors.lastname: Pal
authors.forename: Katerina D
authors.initials: KD
authors.lastname: Kourentzi
authors.forename: Richard C
authors.initials: RC
authors.lastname: Willson
authors.forename: Emery H
authors.initials: EH
authors.lastname: Bresnick
authors.forename: Nader
authors.initials: N
authors.lastname: Sheibani
authors.forename: Samuel H
authors.initials: SH
authors.lastname: Gellman
mesh_terms: D000595:Amino Acid Sequence; D005454:Fluorescence Polarization / Q000379:methods*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D042461:Vascular Endothelial Growth Factor A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D008024:Ligands; D010446:Peptide Fragments; D042461:Vascular Endothelial Growth Factor A