Purification and properties of type 1 topoisomerase from chicken erythrocytes: mechanism of eukaryotic topoisomerase action

Abstract

A simple method for the purification of the major topoisomerase (topoisomerase 1) from chicken erythrocytes is described. Because of the generally repressed state of the chromatin from these nuclei, the heterogeneity of the non-histone proteins is reduced, and it is possible to purify this enzyme from a nuclear extract by a single chromatographic step. The chicken erythrocyte topoisomerase appears to be similar to previously described eukaryotic type I topoisomerases with respect to its physical and enzymological properties. The pattern of intermediate products generated during the action of chicken erythrocyte topoisomerase on a supercoiled closed circular plasmid pBR322 DNA substrate was examined quantitatively and was consistent with a mechanism in which the enzyme closes its substrate DNA molecule after the removal of each superhelical turn and in which dissociation of the enzyme substrate complex may, but does not necessarily, occur after each cycle.