Inhibition of calf thymus type II DNA topoisomerase by poly(ADP-ribosylation).
The effect of poly(ADP-ribosylation) on calf thymus topoisomerase type II reactions has been investigated. Unknotting of phage P4 head DNA, and relaxation and catenation of supercoiled PM2 DNA are inhibited. We conclude that the inhibition results from poly(ADP-ribosylation) on the following grounds. Firstly, the enzyme poly(ADP-ribose) (PADPR) synthetase and NAD are required, secondly, the competitive synthetase inhibitor nicotinamide abolishes topoisomerase inhibition, and thirdly, the polymer alone is not inhibitory. The mechanism of inhibition appears to be disruption of the strand cleavage reaction. A topoisomerase-DNA complex can be formed that upon treatment with protein denaturant at low ionic strength results in strand cleavage. The amount of DNA present in such a cleavable-complex progressively decreased following pretreatment of topoisomerase type II with PADPR synthetase and increasing concentrations of NAD. Treatment of the pre-formed complex with NAD and PADPR synthetase had no effect on its salt-induced dissociation. This suggests that either poly(ADP-ribosylation) has no influence on dissociation of topoisomerase, in contrast to association, or topoisomerase is not accessible to the synthetase when bound to DNA. Similar data were obtained with calf thymus type I topoisomerase.
D000818:Animals; D002417:Cattle; D004247:DNA / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D007700:Kinetics; D009243:NAD / Q000378:metabolism; D009702:Nucleoside Diphosphate Sugars / Q000096:biosynthesis*; D011064:Poly Adenosine Diphosphate Ribose / Q000096:biosynthesis*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D013950:Thymus Gland / Q000201:enzymology*; D059005:Topoisomerase II Inhibitors
D009702:Nucleoside Diphosphate Sugars; D059005:Topoisomerase II Inhibitors; D009243:NAD; D011064:Poly Adenosine Diphosphate Ribose; D004247:DNA; D011065:Poly(ADP-ribose) Polymerases; D004264:DNA Topoisomerases, Type I