Interaction of clinically important human DNA topoisomerase I poison, topotecan, with double-stranded DNA.
Topotecan (TPT), a water-soluble derivative of camptothecin, is a potent antitumor poison of human DNA topoisomerase I (top1) that stabilizes the cleavage complex between the enzyme and DNA. The role of the recently discovered TPT affinity to DNA remains to be defined. The aim of this work is to clarify the molecular mechanisms of the TPT-DNA interaction and to propose the models of TPT-DNA complexes in solution in the absence of top1. It is shown that TPT molecules form dimers with a dimerization constant of (4.0 +/- 0.7) x 10(3) M(-1) and the presence of DNA provokes more than a 400-fold increase of the effective dimerization constant. Flow linear dichroism spectroscopy accompanied by circular dichroism, fluorescence, and surface-enhanced Raman scattering experiments provide evidence that TPT dimers are able to bind DNA by bridging different DNA molecules or distant DNA structural domains. This effect may provoke modification of the intrinsic geometry of the cruciform DNA structures, leading to the appearance of new crossover points that serve as the sites of the top1 loading position. The data presume the hypothesis of TPT-mediated modulation of top1-DNA recognition before ternary complex formation.
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Ul. Vavilova 32, 119991 Moscow, Russia.
Sergei
S
Streltsov
Vladimir
V
Oleinikov
Mikhail
M
Ermishov
Konstantin
K
Mochalov
Alyona
A
Sukhanova
Yuri
Y
Nechipurenko
Sergei
S
Grokhovsky
Alexei
A
Zhuze
Michel
M
Pluot
Igor
I
Nabiev
D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D019281:Dimerization; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D015394:Molecular Structure; D012996:Solutions; D013059:Spectrum Analysis, Raman; D059004:Topoisomerase I Inhibitors; D019772:Topotecan / Q000737:chemistry / Q000378:metabolism*
D004791:Enzyme Inhibitors; D012996:Solutions; D059004:Topoisomerase I Inhibitors; D019772:Topotecan; D004247:DNA