The apurinic/apyrimidinic endodeoxyribonuclease of rat-liver chromatin

Abstract

The extract from rat liver chromatin contains two apurinic/apyrimidinic (AP) endodeoxyribonucleases named 0.2 M and 0.3 M isozymes according to the phosphate concentration necessary to elute them from an hydroxyapatite column. The 0.3 M isozyme is the main and perhaps the only chromatin AP endodeoxyribonuclease in the living cell. This 0.3 M isozyme was purified by successive chromatographies on hydroxyapatite, phosphocellulose, heparin-Sepharose and alkylated-depurinated DNA-cellulose. It has a molecular weight of approximately 39000; its optimum pH is around 8.0; it needs Mg2+ or Mn2+ to be active and the optimum concentration for Mg2+ is between 5 mM and 10 mM. The 0.3 M isozyme has no action on intact DNA strands or on alkylated sites; it cuts the phosphodiester bridge which is the immediate neighbour of the AP site on its 5' side leaving 3'-hydroxyl and 5'-phosphate ends. It has no associated exonuclease activity. To hydrolyze the phosphoester bond near the AP site, the enzyme makes a close contact with three base residues in the large groove of the DNA molecule.