Unit

UNIT 3.3 Topoisomerase Assays

  1. John L. Nitiss1,2,
  2. Eroica Soans1,
  3. Anna Rogojina1,2,
  4. Aman Seth1,
  5. Margarita Mishina1

Published Online: 1 JUN 2012

DOI: 10.1002/0471141755.ph0303s57

Current Protocols in Pharmacology

Current Protocols in Pharmacology

How to Cite

Nitiss, J. L., Soans, E., Rogojina, A., Seth, A. and Mishina, M. 2012. Topoisomerase Assays. Current Protocols in Pharmacology. 57:3.3:3.3.1–3.3.27.

Author Information

  1. 1

    Molecular Pharmacology Department, St. Jude Children's Research Hospital, Memphis, Tennessee

  2. 2

    Biopharmaceutical Sciences Department, University of Illinois College of Pharmacy at Rockford, Rockford, Illinois

Publication History

  1. Published Online: 1 JUN 2012
  2. Published Print: JUN 2012

Abstract

Topoisomerases are nuclear enzymes that play essential roles in DNA replication, transcription, chromosome segregation, and recombination. All cells have two major forms of topoisomerases: type I enzymes, which make single-stranded cuts in DNA, and type II enzymes, which cut and pass double-stranded DNA. DNA topoisomerases are important targets of approved and experimental anti-cancer agents. The protocols described in this unit are for assays used to assess new chemical entities for their ability to inhibit both forms of DNA topoisomerase. Included are an in vitro assay for topoisomerase I activity based on relaxation of supercoiled DNA, and an assay for topoisomerase II based on the decatenation of double-stranded DNA. The preparation of mammalian cell extracts for assaying topoisomerase activity is described, along with a protocol for an ICE assay to examine topoisomerase covalent complexes in vivo, and an assay for measuring DNA cleavage in vitro. Curr. Protoc. Pharmacol. 57:3.3.1-3.3.27. © 2012 by John Wiley & Sons, Inc.

Keywords:

  • topoisomerase;
  • topoisomerase I;
  • topoisomerase II;
  • camptothecin;
  • etoposide;
  • topoisomerase poison