Electrochemical evaluation of Abelson tyrosine-protein kinase 1 activity and inhibition by imatinib mesylate and danusertib

Abstract

Abelson tyrosine-protein kinase 1 (ABL1) catalysed phosphorylation involves the addition of a phosphate group from ATP to the tyrosine residue on the substrate abltide. The phosphorylation reactions were carried out by incubating ABL1, ATP and the substrate abltide. Adsorption at the glassy carbon electrode surface in either reaction mixtures or control solutions, followed by differential pulse voltammetry in buffer allowed detection of the variation of abltide tyrosine residue oxidation peak reflecting the occurrence of the phosphorylation reaction. The effect of abltide, ATP and ABL1 concentrations as well as the time course of the phosphorylation reaction were studied. The influence of co-adsorption of ABL1, ATP and phosphorylated abltide was evaluated and the conditions for the electrochemical detection of ABL1-catalysed phosphorylation optimised. The Michaelis-Menten constant for abltide binding KM∼4.5 μM, turnover number kcat∼11 s(-1) and enzyme efficiency kcat/KM∼2.3 s(-1) μM(-1) were calculated. The inhibition of ABL1 by imatinib mesylate and danusertib was also electrochemically investigated and IC50 values of 0.53 and 0.08 μM determined.

Keywords: Abelson tyrosine-protein kinase 1; Electrochemistry; Enzyme kinetics; Glassy carbon electrode; Inhibition.