Purification and characterization of wheat germ DNA topoisomerase I (nicking-closing enzyme).
Wheat germ contains an enzyme capable of removing supercoils from circular DNA. We have purified this enzyme using Polymin P fractionation, ammonium sulfate precipitation, and chromatography on Bio-Rex 70 and phenyl-Sepharose. Renaturation after electrophoresis on sodium dodecyl sulfate-polyacrylamide gels shows that topoisomerase activity is associated with a polypeptide with a Mr = about 111,000. The enzyme is similar to other eukaryotic type I DNA topoisomerases (nicking-closing enzymes) by the following criteria: it is capable of increasing or decreasing the topological linking number of covalently closed DNA substrate; it is capable of restoring an equilibrium distribution of linking numbers to DNA substrate with a single unique linking number; and it does not require magnesium ion or ATP for activity.
D002413:Cations, Divalent; D002414:Cations, Monovalent; D004264:DNA Topoisomerases, Type I / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D012639:Seeds / Q000201:enzymology*; D014908:Triticum / Q000201:enzymology
D002413:Cations, Divalent; D002414:Cations, Monovalent; D046911:Macromolecular Substances; D004264:DNA Topoisomerases, Type I