title: Mutations of human DNA topoisomerase I at poly(ADP-ribose) binding sites: modulation of camptothecin activity by ADP-ribose polymers.
abstract: 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.
authors.forename: Cinzia
authors.initials: C
authors.lastname: Tesauro
authors.forename: Grazia
authors.initials: G
authors.lastname: Graziani
authors.forename: Barbara
authors.initials: B
authors.lastname: Arnò
authors.forename: Laura
authors.initials: L
authors.lastname: Zuccaro
authors.forename: Alessia
authors.initials: A
authors.lastname: Muzi
authors.forename: Ilda
authors.initials: I
authors.lastname: D'Annessa
authors.forename: Elettra
authors.initials: E
authors.lastname: Santori
authors.forename: Lucio
authors.initials: L
authors.lastname: Tentori
authors.forename: Carlo
authors.initials: C
authors.lastname: Leonetti
authors.forename: Paola
authors.initials: P
authors.lastname: Fiorani
authors.forename: Alessandro
authors.initials: A
authors.lastname: Desideri
mesh_terms: D000595:Amino Acid Sequence; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D001665:Binding Sites; D002166:Camptothecin / Q000494:pharmacology; D004264:DNA Topoisomerases, Type I / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011064:Poly Adenosine Diphosphate Ribose / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D059004:Topoisomerase I Inhibitors / Q000494:pharmacology
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D059004:Topoisomerase I Inhibitors; D011064:Poly Adenosine Diphosphate Ribose; D004264:DNA Topoisomerases, Type I; C528909:TOP1 protein, human; D002166:Camptothecin