9] Escherichia coli phage T4 topoisomerase

Abstract

Topoisomerases are enzymes that alter DNA topology by changing the linking number of circular duplex DNA molecules and by interconverting topologically knotted or catenated DNA forms. The so-called type II topoisomerases act by a mechanism involving the passage of a duplex segment of DNA through a transient double-strand break in another segment of DNA. A novel ATP-dependent type II topoisomerase with DNA-dependent ATPase activity is isolated from extracts of bacteriophage T4-infected E. coli cells. It has a high specific activity for topoisomerization reactions and can be easily purified to near homogeneity in milligram amounts. This chapter explains the interaction of T4 topoisomerase with DNA in the origin region of the T4 chromosome and attempts to reconstitute the initiation reaction in vitro in order to test this and other models for the involvement of the enzyme in the DNA replication process.