Purification and Properties of Nuclear and Cytoplasmic Deoxyribonucleic Acid Polymerases from Human KB Cells

Abstract

We describe the properties of three DNA polymerases isolated from nuclear and cytoplasmic fractions of freshly harvested KB cells. Two of the enzymes, cytoplasmic polylymerase and nuclear polymerase N1, have been highly purified, one of them to apparent homogeneity. The third enzyme, nuclear polymerase N2, which is only partially purified, resembles the cytoplasmic enzyme in its physical properties but possesses distinctive enzymatic characteristics that suggest it may represent an independent third protein. The KB enzymes initiate polymerization at 3′-hydroxyl termini and resemble Escherichia coli DNA polymerase II in their obligatory requirement for gap-containing templates. None of the enzymes can copy the ribonucleotide strand of hybrid templates, even under specific conditions in which E. coli DNA polymerase I does demonstrate such activity. They can, however, with low efficiency copy a deoxynucleotide strand using an oligoribonucleotide as primer. Elucidation of the possible in vivo roles of these enzymes in DNA replication, recombination, and repair must await further biochemical and genetic studies.