Thermodynamics of the multi-stage DNA lesion recognition and repair by formamidopyrimidine-DNA glycosylase using pyrrolocytosine fluorescence--stopped-flow pre-steady-state kinetics.

Nucleic Acids Research
Nikita A KuznetsovOlga S Fedorova

Abstract

Formamidopyrimidine-DNA glycosylase, Fpg protein from Escherichia coli, initiates base excision repair in DNA by removing a wide variety of oxidized lesions. In this study, we perform thermodynamic analysis of the multi-stage interaction of Fpg with specific DNA-substrates containing 7,8-dihydro-8-oxoguanosine (oxoG), or tetrahydrofuran (THF, an uncleavable abasic site analog) and non-specific (G) DNA-ligand based on stopped-flow kinetic data. Pyrrolocytosine, highly fluorescent analog of the natural nucleobase cytosine, is used to record multi-stage DNA lesion recognition and repair kinetics over a temperature range (10-30°C). The kinetic data were used to obtain the standard Gibbs energy, enthalpy and entropy of the specific stages using van't Hoff approach. The data suggest that not only enthalpy-driven exothermic oxoG recognition, but also the desolvation-accompanied entropy-driven enzyme-substrate complex adjustment into the catalytically active state play equally important roles in the overall process.

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Citations

Jun 10, 2014·PloS One·Nikita A KuznetsovOlga S Fedorova
May 7, 2014·Proceedings of the National Academy of Sciences of the United States of America·Shane R NelsonSusan S Wallace
Oct 15, 2013·Environmental and Molecular Mutagenesis·Susan S Wallace
Mar 15, 2015·Biochimica Et Biophysica Acta·Elena S DyakonovaOlga S Fedorova
Dec 19, 2014·Nucleic Acids Research·Nikita A KuznetsovCarlos Simmerling
Nov 8, 2016·Radiation Physics and Chemistry·Andrea J Lee, Susan S Wallace
Jun 3, 2016·Physical Chemistry Chemical Physics : PCCP·Quynh L NguyenSpiridoula Matsika
Oct 1, 2016·Journal of Biomolecular Structure & Dynamics·Nikita A KuznetsovOlga I Lavrik
Aug 21, 2013·The Journal of Biological Chemistry·Maria V LukinaDmitry O Zharkov
Dec 19, 2019·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Lyubov Yu KanazhevskayaOlga S Fedorova
Apr 15, 2015·The Journal of Biological Chemistry·Nikita A KuznetsovOlga S Fedorova
Dec 3, 2016·Biochemistry. Biokhimii︠a︡·N A Kuznetsov, O S Fedorova
Nov 16, 2014·The Journal of Biological Chemistry·Nikita A KuznetsovTatyana V Demidkina
Dec 18, 2020·Frontiers in Cell and Developmental Biology·Alexandra A KuznetsovaNikita A Kuznetsov
Apr 13, 2021·Frontiers in Cell and Developmental Biology·Anastasiia T DavletgildeevaNikita A Kuznetsov
Aug 28, 2021·International Journal of Molecular Sciences·Alexandra A KuznetsovaNikita A Kuznetsov

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Methods Mentioned

BETA
glycosylases
electrophoresis
X-ray

Software Mentioned

DynaFit
Pro Analyzer
Gel

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