STUDIES OF TOPOISOMERASE II AS TARGET OF ACTION OF ANTICANCER DRUG

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The effects of DNA intercalating agents and epipodophyllotoxins are being studied upon purified mammalian topoisomerase II. DNA intercalators and epipodophyllotoxins inhibit DNA topoisomerase II while trapping the enzyme within topoisomerase II-DNA DNA cleavable complexes consit of two enzyme subunits which are covalently bound to the 5'-termini of the DNA break. The DNA breaks can be detected only after protein denaturation by sodium dodecyl sulfate and proteinase K digestion. The breaks also reverse upon salt addition in the absence of added nucleotide. The cleavable complexes trapped by intercalators and epipodophyllotoxins are analogous to intermediates in the DNA breaking-rejoining reaction of topoisomerase II. It is by this reaction that the enzyme carries out DNA strand passage and DNA topoisomerization reaction. Another mode of interaction between topoisomerase II-DNA complexes and intercalator is the forced reversal or unlocking of cleavable complexes. This effect has been observed with high concentrations of 2-methyl-9-hydorxyellipticinium and with the bisintercalator, ditercalinium. The unlocking of cleavable complexes seems to be related to DNA binding and may not be accompanied by enzyme inhibition. The localization of intercalator-induced topoisomerase II action in the nuclear genome was investigated. It was found that certain repeated sequences are enriched at a favored distance from sites of intercalator-induced topoisomerase II-DNA linkage. The results have implications regarding the localization of topoisomerase II action in the loop structure of nuclear chromatin.