Topoisomerase II site-directed alkylation of DNA by psorospermin and its effect on topoisomerase II-mediated DNA cleavage

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33 Scopus citations

Abstract

Psorospermin, a plant-derived antitumor agent, has been shown to selectively alkylate a guanine at the topoisomerase II cleavage site to trap the topoisomerase II-DNA cleaved complex. The results of this study provide further important insight into the mechanism of the topoisomerase II site- directed alkylation of DNA by psorospermin and its subsequent effects on the topoisomerase II-induced DNA cleavage. First, we demonstrate that the topoisomerase II-induced alkylation of DNA by psorospermin occurs at a time preceding the topoisomerase II-mediated strand cleavage event, because it occurs in the absence of Mg2+. We confirm that the alkylation of DNA by psorospermin takes place at N-7 of guanine in the presence of topoisomerase II, because substitution of the target guanine by 7-deazaguanine prevents alkylation. Because the stimulation of the topoisomerase II-induced DNA cleavage by psorospermin can be slowly reversed by the addition of excess salt, this indicates that alkylation of DNA by psorospermin traps a reversible topoisomerase II-DNA complex. Both the DNA alkylation by psorospermin in the presence of topoisomerase II and the enzyme-mediated DNA cleavage elevated by psorospermin are more enhanced at acidic pH values, in accordance with the increased stability of the topoisomerase II-DNA complex at acidic pH values. Finally, our results suggest that it is the psorospermin-DNA adducts, not the abasic sites resulting from depurination, that are responsible for the stimulation of the topoisomerase II-mediated cleavage. Because the precise location of the psorospermin within the topoisomerase II cleavage site is known, together with the covalent DNA linkage chemistry and the conformation of the psorospermin-DNA adduct, this structural insight provides an excellent opportunity for the design and synthesis of new, more effective topoisomerase II poisons.

Original languageEnglish (US)
Pages (from-to)33020-33026
Number of pages7
JournalJournal of Biological Chemistry
Volume273
Issue number49
DOIs
StatePublished - Dec 4 1998
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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