HUMAN 170-KDA AND 180-KDA TOPOISOMERASES-II BIND PREFERENTIALLY TO CURVED AND LEFT-HANDED LINEAR DNA

The binding activities of the 170 kDa and the 180 kDa human topoisomerases II (topo II alpha and topo II beta) to linear DNA fragments with different degrees of curvature were characterized. In gel retardation experiments it was shown that both forms of the enzyme bind preferentially to a curved 287 bp fragment, forming a detectable stable complex. The affinity for straight DNA fragments of similar length is significantly lower, Both a commercially available topo II alpha, isolated from placenta, and topo II alpha and topo II beta purified from nuclear extracts of the Namalwa lymphoma tissue culture line gave similar results. The effects of double-stranded poly[d(A-T)], poly[d(G-C)], supercoiled plasmid DNA and linear Z-DNA on the topo II-complex with curved DNA were analyzed in competition experiments. The hierarchy of affinities of the 180 kDa topo II beta for these DNAs has the order: linear left-handed DNA > supercoiled DNA greater than or equal to curved DNA much greater than poly[d(A-T)] > poly[d(G-C)]. The 170 kDa topo II alpha binds with similar affinity to curved DNA and linear Z-DNA greater than or equal to supercoiled DNA much greater than linear B-DNA The data imply that human topoisomerase II binding is more sensitive to DNA secondary structure than to DNA sequence per se. The ability of the enzyme to preferentially recognize a wide variety of sequences in unusual secondary structures suggests a mode of targeting the enzyme in vivo to regions of high negative supercoiling.