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Human apurinic/apyrimidinic endonuclease is processive
1Department of Biology, Northeastern University, Boston, Massachusetts 02115, USA.
Biochemistry
|December 22, 1999
Summary
Apurinic/apyrimidinic endonuclease (AP endo) repairs oxidative DNA damage by nicking abasic sites. This study shows AP endo acts quasi-processively, cleaving multiple sites before dissociation, suggesting regulated DNA repair mechanisms.
Area of Science:
- Molecular Biology
- DNA Repair Mechanisms
- Enzymology
Background:
- Apurinic/apyrimidinic endonuclease (AP endo) is crucial for base excision repair of oxidative DNA damage.
- AP endo initiates repair by nicking double-stranded DNA 5' to abasic sites.
Purpose of the Study:
- To investigate whether AP endo locates abasic sites via a processive or distributive mechanism.
- To characterize the processivity and catalytic activity of AP endo on multi-abasic site substrates.
Main Methods:
- Synthesis of a linear multi-abasic site DNA substrate (concatemer) from 25-nucleotide monomers.
- Enzymatic assays using the concatemer to determine AP endo's cleavage activity and processivity.
- Analysis of steady-state and single-turnover kinetic parameters.
Main Results:
- AP endo exhibited similar nicking activity on monomeric and concatemeric substrates.
- AP endo cleaved approximately 7-8 abasic sites, traveling over 200 nucleotides, indicating quasi-processive behavior.
- Processivity and catalytic rate were separable and differentially regulated by salt concentration, with optimal processivity at intermediate salt levels.
Conclusions:
- AP endo functions in a quasi-processive manner, similar to uracil DNA glycosylase.
- The processivity of AP endo is subject to regulation by ionic strength, potentially fine-tuning DNA repair at physiological salt concentrations.