Net DNA synthesis catalysed by calf thymus DNA polymerase beta.

ABSTRACT

1. The amounts of deoxynucleotides incorporated during an extensive replication by DNA polymerase alpha into poly(dA)-oligo(dT)12-18 and damaged DNA containing 2.5 incisions per molecule and 3.7% single-stranded DNA, corresponded to the amounts of non-complexed poly(dA) and single-stranded fraction of DNA, respectively. The amounts of the corresponding DNA polymerase beta products were several times higher. In the case of activated DNA they exceeded input DNA. The DNA polymerase beta reaction on this template was continued till substrate exhaustion. 2. The reaction of DNA polymerase beta with activated DNA, leading to net DNA synthesis, was template-directed, required Mg2+ and four deoxynucleoside triphosphates; was not inhibited by DNA polymerase alpha inhibitors, but was sensitive to 2',3'-dideoxythymidine triphosphate. The DNA product was completely digestable by DNAse I and showed a base ratio typical of calf thymus DNA. 3. The essential difference in the reaction mechanism between DNA polymerase alpha and beta suggests the ability of the latter enzyme to synthesize DNA with displacement of the non-replicated strand.