title: Gaps in DNA induced by neocarzinostatin bear 3'- and 5'-phosphoryl termini.
abstract: Neocarzionstatin (NCS)-induced strand breakage of DNA generates nonfunctional binding sites for the E. coli DNA polymerase I. Treatment of the NCS-nicked DNA with alkaline phosphatase at 65 degrees C prior to the polymerase reaction results in 60-100-fold stimulation of dTMP incorporation whereas in a control not treated with the drug there is only a 2-fold increase. Sites of strand scission on the NCS-treated DNA bear phosphate at the 3' termini. This conclusion is supported by the kinetics of release of inorganic phosphate from NCS-cut DNA by exonuclease III. Since our earlier work has shown that virtually all the 5' ends of the nicks caused by NCS bear phosphomonoester groupings, the 3'- and 5'- phosphoryl termini could be quantitated using alkaline phosphatase and exonuclease III. Over a wide range of drug levels the amount of inorganic phosphate released by alkaline phosphatase is approximately twice as much as that removed by exonuclease III, indicating the presence of equal amounts of 3'- and 5'- phosphoryl termini. This, taken together with other previously demonstrated effects of NCS on DNA, such as the introduction of nicks not sealable by polynucleotide ligase, the release of thymine, and the formation of a malonaldehyde type compound, suggests that NCS-induced strand breakage involves base release accompanied by opening of the sugar ring with destruction of one or more nucleosides and results in a gap bounded by 3'- and 5'- phosphoryl termini.
mesh_terms: D000469:Alkaline Phosphatase; D000903:Antibiotics, Antineoplastic; D055598:Chemical Phenomena; D002621:Chemistry; D004247:DNA; D004256:DNA Polymerase I / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D005092:Exonucleases; D007700:Kinetics; D013379:Substrate Specificity; D013698:Templates, Genetic; D009353:Zinostatin
chemical_list: D000903:Antibiotics, Antineoplastic; D004247:DNA; D009353:Zinostatin; D004256:DNA Polymerase I; D005092:Exonucleases; D000469:Alkaline Phosphatase