Mutations of human DNA topoisomerase I at poly (ADP-ribose) binding sites: Modulation of camptothecin activity by ADP-ribose polymers

Background: dna topoisomerases are key enzymes that modulate the topological state of dna through the breaking and rejoining of dna strands. human topoisomerase i belongs to the family of poly(adp-ribose)-binding proteins and is the target of camptothecin derived anticancer drugs. poly(adp-ribosyl)ation occurs at specific sites of the enzyme inhibiting the cleavage and enhancing the religation steps during the catalytic cycle. thus,adp-ribose polymers antagonize the activity of topoisomerase i poisons,whereas parp inhibitors increase their antitumor effects. methods: using site-directed mutagenesis we have analyzed the interaction of human topoisomerase i and poly (adp-ribose) through enzymatic activity and binding procedures. results: mutations of the human topoisomerase i hydrophobic or charged residues,located on the putative polymer binding sites,are not sufficient to abolish or reduce the binding of the poly(adp-ribose) to the protein. these results suggest either the presence of additional binding sites or that the mutations are not enough perturbative to destroy the poly(adp-ribose) interaction,although in one mutant they fully abolish the enzyme activity. conclusions: it can be concluded that mutations at the hydrophobic or charged residues of the putative polymer binding sites do not interfere with the ability of poly(adp-ribose) to antagonize the antitumor activity of topoisomerase i poisons. © 2014 tesauro et al.; licensee biomed central ltd.