Description | E.coliTopoisomeraseIhasalsobeenimplicatedinknottingandunknottingDNAandinlinkingcomplementaryringsofsingle-strandedDNAintodouble-strandedrings.Theintactholoenzymeisa97KproteinwiththreeZn(II)atomsintertacysteinemotifsnearitscarboxy-terminus.TopoisomeraseIappearstoreversesupercoilingbytransientlybreakingasegmentofsingle-strandedDNA,passinganintactsingle-ordouble-strandedstrandofDNAthroughthegate,thenrejoiningthebrokensegment. |
Source | AnE.colistrainthatcarriestheE.colitopAgene |
UnitDefinition | Oneunitisdefinedastheamountofenzymethatcatalyzestherelaxationof>95%of0.5μgofpUC19RFI(negativelysupercoiled)DNAin15minutesat37°C. |
Components | E.coliTopoisomeraseI:5,000units/mlin10mMTris-HCl,1mMDTT,0.1mMEDTA,50%glycerol,pH7.5@25°C |
QualityControl | Theabsenceofexo-andendodeoxyribonucleases,non-specificDNaseandRNaseisconfirmedbyappropriatequalitytest. |
StorageCondition | -20°C |