title: Kinetic mechanism of the interaction of Saccharomyces cerevisiae AP-endonuclease 1 with DNA substrates.
abstract: The apurinic/apyrimidinic endonuclease from Saccharomyces cerevisiae Apn1 is one of the key enzymes involved in base excision repair of DNA lesions. A major function of the enzyme is to cleave the upstream phosphodiester bond of an apurinic/apyrimidinic site (AP-site), leading to the formation of a single-strand break with 3'-hydroxyl (OH) and 5'-deoxyribose phosphate (dRP) termini. In this study, the pre-steady-state kinetics and conformational dynamics of DNA substrates during their interaction with Apn1 were investigated. A stopped-flow method with detection of the fluorescence intensity of 2-aminopurine and pyrrolocytosine located adjacent or opposite to the damage was used. It was found that upon interaction with Apn1, both DNA strands undergo a number of rapid changes. The location of fluorescent analogs of heterocyclic bases in DNA does not influence the catalytic step of the reaction. Comparison of data obtained for yeast Apn1 and reported data (Kanazhevskaya, L. Yu., Koval, V. V., Vorobjev, Yu. N., and Fedorova, O. S. (2012) Biochemistry, 51, 1306-1321) for human Ape1 revealed some differences in their interaction with DNA substrates.
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, 630090 Novosibirsk, Russia.
authors.forename: E S
authors.initials: ES
authors.lastname: Dyakonova
authors.forename: V V
authors.initials: VV
authors.lastname: Koval
authors.forename: A A
authors.initials: AA
authors.lastname: Ishchenko
authors.forename: M K
authors.initials: MK
authors.lastname: Saparbaev
authors.forename: R
authors.initials: R
authors.lastname: Kaptein
authors.forename: O S
authors.initials: OS
authors.lastname: Fedorova
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D045643:DNA Repair Enzymes / Q000378:metabolism*; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D004706:Endodeoxyribonucleases; C083274:Apn1 protein, S cerevisiae; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D045643:DNA Repair Enzymes