Structural insight into a quinolone-topoisomerase II-DNA complex - Furtherevidence for a 2 : 2 quinobenzoxazine-Mg2+ self-assembly model formed in the presence of topoisomerase II

Quinobenzoxazine A-62176, developed from the antibacterial fluoroquinolones , is active in vitro and in vivo against murine and human tumors. It has be en previously claimed that A-62176 is a catalytic inhibitor of mammalian to poisomerase II that does not stabilize the cleaved complex. However, at low drug concentrations and pH 6-7, we have found that A-62176 can enhance the formation of the cleaved complex at certain sites. Using a photocleavage a ssay, mismatched sequences, and competition experiments between psorospermi n and A-62176, we pinpointed the drug binding site on the DNA base pairs be tween positions +1 and +2 relative to the cleaved phosphodiester bonds. A 2 :2 quinobenzoxazine-Mg2+ self-assembly model was previously proposed, in wh ich one drug molecule intercalates into the DNA helix and the second drug m olecule is externally bound, held to the first molecule and DNA by two Mg2 bridges, The results of competition experiments between psorospermin and A -62176, as well as between psorospermin and A-62176 and norfloxacin, are co nsistent with this model and provide the first evidence that this 2:2 quino benzoxazine-Mg2+ complex is assembled in the presence of topoisomerase II. These results also have parallel implications for the mode of binding of th e quinolone antibiotics to the bacterial gyrase-DNA complex.