An inhibitor of DNA topoisomerase I from Xenopus laevis ovaries

Abstract

A novel, heat-resistant and Pronase-sensitive, inhibitor of eukaryotic DNA topoisomerase I has been purified from Xenopus laevis ovaries. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the most purified fraction revealed three bands with apparent molecular masses of 25, 28.5, and 33.5 kDa. The 25- and 33.5-kDa peptides recovered from an SDS-PAGE gel inhibited X. laevis DNA topoisomerase I. The purified inhibitor was specific to DNA topoisomerase I and did not inhibit other DNA enzymes tested. The inhibitor blocked the catalytic activity of DNA topoisomerase I by interacting with the enzyme, rather than by competing for binding sites on substrate DNA. Binding of DNA topoisomerase I to substrate DNA was blocked by the inhibitor, as was the cleavage reaction catalyzed by DNA topoisomerase I. Inhibition of DNA topoisomerase I was relieved by divalent cations Ca2+, Mg2+, or Mn2+.

Casein Kinase II Stimulates Xenopus laevis DNA Topoisomerase I by Physical Association

“…Zhao and Benbow have recently identified and purified to homogeneity a X. laevis ovarian protein that inhibits X. laevis DNA topoisomerase I activity, apparently by also interacting directly with the enzyme (Zhao & Benbow, 1993). Taken together with the results of this study showing direct stimulation of X .…”

“…Zhao and Benbow have recently identified and purified to homogeneity a X. laevis ovarian protein that inhibits X. laevis DNA topoisomerase I activity, apparently by also interacting directly with the enzyme (Zhao & Benbow, 1993). Taken together with the results of this study showing direct stimulation of X. laevis DNA topoisomerase I by physical association with casein kinase II, these findings suggest that direct physical association and the accompanying proteinprotein interactions may play a larger role than previously envisioned in the regulation of DNA topoisomerase I activity.…”