THE INHIBITORY EFFECT OF NOVEL TRITERPENOID COMPOUNDS, FOMITELLIC ACIDS, ON DNA-POLYMERASE-BETA

We previously found new triterpenoid compounds, designated fomitellic acid A and B, which selectively inhibit the activities of mammalian DN A polymerase alpha and beta in vitro. On DNA polymerase beta, the fomi tellic acids acted by competing with both the substrate and the templa te primer, but on DNA polymerase alpha, they acted non-competitively. At least on DNA polymerase beta, the evidence suggests that each of th e fomitellic acids bind to the active region competing with the substr ate and/or template primer, and subsequently inhibits the catalytic ac tivity. We therefore further investigated the enzyme-binding propertie s by using DNA polymerase beta and its proteolytic fragments. The 39 k Da enzyme was proteolytically separated into two fragments of the temp late-primer-binding domain (8 kDa) and the catalytic domain (31 kDa). The fomitellic acids bound tightly to the 8 kDa fragment, but not to t he 31 kDa fragment. The immunoprecipitation by antibodies against the enzyme or each of the fragments also proved the binding. These results suggest that the fomitellic acid molecule competes with the template- primer molecule on its 8 kDa binding site, binds to the site, and the fomitellic acid molecule simultaneously disturbs the substrate incorpo ration into the template primer.