Ah. Sarker et al., PURIFICATION AND CHARACTERIZATION OF AN AP ENDONUCLEASE DNA 3'REPAIR DIESTERASE FROM MOUSE ASCITES SARCOMA-CELLS, Biochimica et biophysica acta (G). General subjects, 1245(3), 1995, pp. 299-304
Purification and characterization of a DNA repair enzyme having 5' apu
rinic/apyrimidinic (AP) endonuclease activity are reported. The enzyme
extracted from mouse ascites sarcoma (SR-C3H/He) cells with 0.2 M pot
assium phosphate buffer (pH 7.5) was purified by successive chromatogr
aphies on phosphocellulose, DEAE-cellulose, phosphocellulose (a second
time) and single-stranded DNA cellulose, and by sodium dodecyl sulfat
e-polyacrylamide gel electrophoresis (SDS-PAGE). The purified enzyme h
as an apparent molecular mass of 30 kDa as determined by SDS-PAGE. It
was shown to have nicking activity on acid-depurinated DNA but not on
intact DNA, and to have priming activities for DNA polymerase on acid-
depurinated DNA and bleomycin-treated DNA. The results indicate that i
t is a multifunctional DNA repair enzyme having 5' AP endonuclease and
DNA 3' repair diesterase activities. The enzyme activity is dependent
upon the presence of a divalent cation such as Mg2+. Its amino-termin
al amino acid acid internal amino acid sequences are determined.