title: Exploring "one-shot" kinetics and small molecule analysis using the ProteOn XPR36 array biosensor.
abstract: A ProteOn XPR36 parallel array biosensor was used to characterize the binding kinetics of a set of small molecule/enzyme interactions. Using one injection with the ProteOn's crisscrossing flow path system, we collected response data for six different concentrations of each analyte over six different target protein surfaces. This "one-shot" approach to kinetic analysis significantly improves throughput while generating high-quality data even for low-molecular-mass analytes. We found that the affinities determined for nine sulfonamide-based inhibitors of the enzyme carbonic anhydrase II were highly correlated with the values determined using isothermal titration calorimetry. We also measured the temperature dependence (from 15 to 35 degrees C) of the kinetics for four of the inhibitor/enzyme interactions. Our results illustrate the potential of this new parallel-processing biosensor to increase the speed of kinetic analysis in drug discovery and expand the applications of real-time protein interaction arrays.
authors.affiliation: Bio-Rad Haifa, Haifa 32000, Israel.
authors.forename: Tsafrir
authors.initials: T
authors.lastname: Bravman
authors.forename: Vered
authors.initials: V
authors.lastname: Bronner
authors.forename: Kobi
authors.initials: K
authors.lastname: Lavie
authors.forename: Ariel
authors.initials: A
authors.lastname: Notcovich
authors.forename: Giuseppe A
authors.initials: GA
authors.lastname: Papalia
authors.forename: David G
authors.initials: DG
authors.lastname: Myszka
mesh_terms: D015374:Biosensing Techniques; D024402:Carbonic Anhydrase II / Q000187:drug effects / Q000378:metabolism; D002257:Carbonic Anhydrase Inhibitors / Q000494:pharmacology; D007700:Kinetics; D008970:Molecular Weight; D011485:Protein Binding; D013449:Sulfonamides / Q000494:pharmacology; D013696:Temperature
chemical_list: D002257:Carbonic Anhydrase Inhibitors; D013449:Sulfonamides; D024402:Carbonic Anhydrase II