title: Thermodynamics of the multi-stage DNA lesion recognition and repair by formamidopyrimidine-DNA glycosylase using pyrrolocytosine fluorescence--stopped-flow pre-steady-state kinetics.
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.
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of Russian Academy of Sciences, Novosibirsk, 630090, Russia.
authors.forename: Nikita A
authors.initials: NA
authors.lastname: Kuznetsov
authors.forename: Yuri N
authors.initials: YN
authors.lastname: Vorobjev
authors.forename: Lev N
authors.initials: LN
authors.lastname: Krasnoperov
authors.forename: Olga S
authors.initials: OS
authors.lastname: Fedorova
mesh_terms: D003596:Cytosine / Q000031:analogs & derivatives* / Q000737:chemistry; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D043382:DNA-Formamidopyrimidine Glycosylase / Q000737:chemistry* / Q000378:metabolism; D005453:Fluorescence; D005663:Furans / Q000378:metabolism; D006151:Guanosine / Q000031:analogs & derivatives / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D013816:Thermodynamics
chemical_list: C576136:3-(b-D-2-ribofuranosyl)-6-methylpyrrolo(2,3-d)pyrimidin-2(3H)-one; D005663:Furans; D006151:Guanosine; C046215:8-hydroxyguanosine; C018674:tetrahydrofuran; D003596:Cytosine; D004247:DNA; D043382:DNA-Formamidopyrimidine Glycosylase