Conformational Dynamics of Dioxygenase AlkB and DNA in the Course of Catalytically Active Enzyme–Substrate Complex Formation

Abstract

Fe²⁺/2-ketoglutarate-dependent DNA-dioxygenase AlkB from Escherichia coli is able to restore the native structure of alkylated DNA bases. The enzymatic process utilizes the molecular oxygen, and proceeds through a mechanism of oxidative dealkylation. Here, the kinetics of conformational changes of AlkB and DNA substrates in the course of binding steps were studied. Nickel and cobalt divalent ions were used instead of Fe²⁺ as metal cofactors in order to inhibit the catalytic activity of AlkB and to study certain stages leading to the formation of a catalytically active enzyme–substrate complex.