Isolation of DNA Topoisomerases I and II Activities from Mitochondria of Mammalian Heart

DNA topoisomerases constitute a ubiquitous class of enzymes that engage chromosomal DNA and promote the passage of the two strands of the DNA helix through one another (reviewed in refs. 1 and 2 ). This occurs while preserving the phosphodiester backbone of each strand. In some instances, the DNA topoisomerase promotes passage of one helical segment of duplex DNA through another segment. The ability to transiently break DNA, pass DNA through the break, and then rejoin the DNA gives the DNA topoisomerases the ability to change the linking number of DNA (the number of times the two strands of DNA in a constrained chromosome revolve around one another). In vivo, the DNA topoisomerase activities are needed to alleviate torsional strain imposed on the DNA helix. They eliminate, for example, the overwinding or underwinding of the DNA helix that occurs during DNA replication, transcription, and at the end of the DNA replication cycle when daughter chromosome helices need to be disentangled from one another. In vitro, this topoisomerase activity is reflected in the enzyme’s capacity to remove supercoils from supercoiled DNA templates, as well as to knot/unknot and catenate/decatenate circular, covalently closed DNA circular substrates.