Journal
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
Volume 1865, Issue 2, Pages -Publisher
ELSEVIER
DOI: 10.1016/j.bbagen.2020.129772
Keywords
Thermodynamic binding data; Kinetic binding data; Measurement protocol; kinITC; 1:1 protein-ligand complexes; Drug optimization
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This study presents an efficient protocol for extracting thermodynamic and kinetic data for 1:1 protein-ligand reactions from AFFINImeter kinITC in a single experiment. The protocol, which requires the same time as the standard one, improves the precision of both thermodynamic and kinetic data compared to the standard protocol.
Background: Thermodynamic and binding kinetic data increasingly support and guide the drug optimization process. Methods: Because ITC thermograms contain binding thermodynamic and kinetic information, an efficient protocol for the simultaneous extraction of thermodynamic and kinetic data for 1:1 protein ligand reactions from AFFINImeter kinITC in one single experiment are presented. Results: The effort to apply this protocol requires the same time as for the standard protocol but increases the precision of both thermodynamic and kinetic data. Conclusions: The protocol enables reliable extraction of both thermodynamic and kinetic data for 1:1 protein-ligand binding reactions with improved precision compared to the 'standard protocol'. General significance: Thermodynamic and kinetic data are recorded under exactly the same conditions in solution without any labeling or immobilization from a protein sample that is not 100% active and would otherwise render the extraction of kinetic parameters impossible.
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