Conformational Dynamics of Dioxygenase AlkB and DNA in the Course of Catalytically Active Enzyme–Substrate Complex Formation
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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.
Keywords:DNA dioxygenase AlkB demethylation of DNA conformational dynamics pre-steady-state kinetics
The authors are grateful to M.K. Saparbaev (Gustave Roussy Institute, Villejuif, France) for the kind gift of the alkB gene.
The work was partially supported by the basic budgetary investment of PFNI GAN 2013–2020, project no. ААА-А17-117020210022-4. Part of the work on pre-steady-state kinetics was supported by the Russian Science Foundation (project no. 16-14-10038).
COMPLIANCE WITH ETHICAL STANDARDS
The work has no studies involving humans or animals as subjects of the study.
Conflict of Interests
The authors declare they have no conflicts of interest.