EFFECTS OF NUCLEOSOMES AND ANTITUMOR DRUGS ON THE CATALYTIC ACTIVITY OF TYPE-II DNA TOPOISOMERASE FROM RAT TESTIS

To gain insight into the relative catalytic efficiencies of mammalian type I and type II DNA topoisomerases, in the cellular context, we hav e used naked DNA and DNA incorporated into nucleosomes as substrates. We observed that the relaxation activity of both the enzymes declined with DNA containing increasing densities of nucleosomes; however, kine tic analysis revealed that topoisomerase I seemed less affected than t opoisomerase II. The addition of histone H1, in stoichiometric amounts , to naked DNA or minichromosomes lessened the activity of topoisomera se II, and required 7-fold less for complete inhibition when the latte r was used as the substrate. To ascertain if the observed differences are specific to topoisomerase II from testis, we examined the effect o f nucleosomes on the catalytic efficiency of its isoform from liver. I nterestingly, the suppression of relaxation activity of Liver topoisom erase II required substrates containing higher mass ratios of histone octamer/DNA. Studies on the effect of nucleosomes an the action of ten iposide displayed significant differences in the kinetics of the react ion, in its IC50 values, and have provided biochemical evidence for th e first time that nucleosomes increased inhibition caused by teniposid e. Further, this feature appears to be specific for topoisomerase II-d irected drugs and is not shared by the generic class of either DNA-int ercalating or non-DNA-intercalating ligands. (C) Elsevier Science Inc.