The noncovalent complex between DNA and the bifunctional intercalator ditercalinium is a substrate for the UvrABC endonuclease of Escherichia coli.
We have demonstrated that the noncovalent complex formed between DNA and an antitumor bifunctional intercalator, ditercalinium, is recognized in vitro as bulky covalent DNA lesions by the purified Escherichia coli UvrABC endonuclease. It was established that no covalent drug-DNA adduct was formed during the incubation of the drug with DNA or during subsequent incubation with the UvrAB proteins. The nucleoprotein-ditercalinium complexes appear different from those generated by repair of pyrimidine dimers. The UvrA protein is able to form a stable complex with ditercalinium-intercalated DNA in the presence of ATP, whereas both UvrA and UvrB proteins are required to form a stable complex with pyrimidine dimer-containing DNA. The apparent half-life of the UvrA- and UvrAB-ditercalinium-DNA complexes following removal of free ditercalinium is 5 min. However, if the free ditercalinium concentration is maintained to allow the intercalation of one molecule of ditercalinium per 3000 base pairs, the half-life of the UvrA- or UvrAB-ditercalinium-DNA complex is 50 min, comparable to that of the complex of UvrAB proteins formed with pyrimidine dimer-containing DNA. UvrABC endonuclease incises ditercalinium-intercalated DNA as efficiently as pyrimidine dimer-containing DNA. However, unlike repair of pyrimidine dimers, the incision reaction is strongly favored by the supercoiling of the DNA substrate. Because UvrA- or UvrAB-ditercalinium-DNA complexes can be formed with relaxed DNA without leading to a subsequent incision reaction, these apparently dead-end nucleoprotein complexes may become lesions in themselves resulting in the cytotoxicity of ditercalinium. Our results show that binding of excision repair proteins to a noncovalent DNA-ligand complex may lead to cell toxicity.
Lambert
B
B
Unité de Physicochimie Macromoléculaire, URA 158 Centre National de la Recherche Scientifique, Institut Gustave Roussy, Villejuif, France.
Jones
B K
BK
Roques
B P
BP
Le Pecq
J B
JB
Yeung
A T
AT
D000970:Antineoplastic Agents / Q000378:metabolism*; D002227:Carbazoles / Q000378:metabolism* / Q000494:pharmacology; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D007364:Intercalating Agents / Q000378:metabolism; D007700:Kinetics; D010957:Plasmids; D011485:Protein Binding; D013379:Substrate Specificity
D000970:Antineoplastic Agents; D002227:Carbazoles; D029968:Escherichia coli Proteins; D007364:Intercalating Agents; D004706:Endodeoxyribonucleases; C036385:endodeoxyribonuclease uvrABC; C042804:ditercalinium