title: Measurement and differentiation of ligand-induced calmodulin conformations by dual polarization interferometry.
abstract: In early drug discovery, knowledge about ligand-induced conformational changes and their influence on protein activity greatly aids the identification of lead candidates for medicinal chemistry efforts. Efficiently acquiring such information remains a challenge in the initial stages of lead finding. Here we investigated the application of dual polarization interferometry (DPI) as a method for the real-time characterization of low molecular weight (LMW) ligands that induce conformational changes. As a model system we chose calmodulin (CaM), which undergoes large and distinct structural rearrangements in response to calcium ion and small molecule inhibitors such as trifluoperazine (TFP). We measured concentration-dependent mass, thickness, and density responses of an immobilized CaM protein layer, which correlated directly with binding and conformational events. Calcium ion binding to CaM induced an increase in thickness (≤0.05 nm) and decrease in density (≤-0.03 g/cm(3)) whereas TFP induced an increase in both thickness (≤0.05 nm) and density (≤0.01 g/cm(3)). The layer measurements reported here show how DPI can be used to assess and differentiate ligands with distinct structural modes of action.
authors.affiliation: Novartis Institutes for BioMedical Research Basel, CPC/LFP/Label-free Technologies, Novartis Pharma AG, Basel, Switzerland. kristin.coan@novartis.com
authors.forename: Kristin E D
authors.initials: KE
authors.lastname: Coan
authors.forename: Marcus J
authors.initials: MJ
authors.lastname: Swann
authors.forename: Johannes
authors.initials: J
authors.lastname: Ottl
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism; D002417:Cattle; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism; D007368:Interferometry; D008024:Ligands; D011485:Protein Binding; D014268:Trifluoperazine / Q000737:chemistry
chemical_list: D002147:Calmodulin; D004800:Enzymes, Immobilized; D008024:Ligands; D014268:Trifluoperazine; D002118:Calcium