REVIEW: Thermodynamic Analysis of Fast Stages of Specific Lesion Recognition by DNA Repair Enzymes

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N. A. Kuznetsov^* and O. S. Fedorova^*

Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, 630090 Novosibirsk, Russia; E-mail: nikita.kuznetsov@niboch.nsc.ru, fedorova@niboch.nsc.ru

^* To whom correspondence should be addressed.

Received May 30, 2016; Revision received June 29, 2016

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The methodology of determination of the thermodynamic parameters of fast stages of recognition and cleavage of DNA substrates is described for the enzymatic processes catalyzed by DNA glycosylases Fpg and hOGG1 and AP endonuclease APE1 during base excision repair (BER) pathway. For this purpose, stopped-flow pre-steady-state kinetic analysis of tryptophan fluorescence intensity changes in proteins and fluorophores in DNA substrates was performed at various temperatures. This approach made it possible to determine the changes of standard Gibbs free energy, enthalpy, and entropy of sequential steps of DNA-substrate binding, as well as activation enthalpy and entropy for the transition complex formation of the catalytic stage. The unified features of mechanism for search and recognition of damaged DNA sites by various enzymes of the BER pathway were discovered.

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KEY WORDS: thermodynamics, pre-steady-state kinetics, DNA glycosylase, AP endonuclease, DNA damage, base excision repair

DOI: 10.1134/S0006297916100114

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