Biochemistry (Moscow)

, Volume 77, Issue 10, pp 1162–1171

Kinetic mechanism of the interaction of Saccharomyces cerevisiae AP-endonuclease 1 with DNA substrates

  • E. S. Dyakonova
  • V. V. Koval
  • A. A. Ishchenko
  • M. K. Saparbaev
  • R. Kaptein
  • O. S. Fedorova
Article

DOI: 10.1134/S0006297912100082

Cite this article as:
Dyakonova, E.S., Koval, V.V., Ishchenko, A.A. et al. Biochemistry Moscow (2012) 77: 1162. doi:10.1134/S0006297912100082
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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.

Key words

Apn1 base excision repair (BER) stopped-flow method 2-aminopurine pyrrolocytosine 

Abbreviations

Ape1

human apurinic/apyrimidinic endonuclease

Apn1

apurinic/apyrimidinic endonuclease from Saccharomyces cerevisiae

AP-site

abasic site

2-aPu

2-aminopurine

BER

base excision repair

F

synthetic analog of AP-site ((3-hydroxytetrahydrofuran-2-yl)methyl phosphate)

PyrC

pyrrolocytosine (3-[β-D-2-ribofuranosyl]-6-methylpyrrolo[2,3-d]pyrimidin-2(3H)-one)

Copyright information

© Pleiades Publishing, Ltd. 2012

Authors and Affiliations

  • E. S. Dyakonova
    • 1
    • 2
  • V. V. Koval
    • 1
    • 2
  • A. A. Ishchenko
    • 3
  • M. K. Saparbaev
    • 3
  • R. Kaptein
    • 2
    • 4
  • O. S. Fedorova
    • 1
    • 2
  1. 1.Institute of Chemical Biology and Fundamental MedicineSiberian Branch of the Russian Academy of SciencesNovosibirskRussia
  2. 2.Novosibirsk State UniversityNovosibirskRussia
  3. 3.UMR 8200 C.N.R.S. Institut Gustave RoussyGroupe “Reparation de l’ADN” Univ. Paris-Sud XIVillejuif CedexFrance
  4. 4.Bijvoet Center for Biomolecular ResearchUtrecht UniversityUtrechtThe Netherlands

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