Methods in Molecular Biology DNA Topoisomerase Protocols

Beginning with the Escherichia coli co protein, or bacterial DNA topoisomerase I, an ever-increasing number of enzymes has been identified that catalyze changes in the linkage of DNA strands. DNA Topoisomerase Protocols, I: DNA Topology and Enzymes brings together an unprecedented collection of cutting-edge experimental protocols for investigating DNA structure, topology, and DNA topoisomerase function. Described by expert experimentalists who have perfected the techniques, these unfailingly reproducible methods utilize new approaches to study changes in DNA topology (linking number, knotting, catenation, and relaxation) and DNA structure (bending), as well as to assess chromosome structure through FLP-mediated recombination and to analyze bacterial nucleoid structure. State-of-the-art protocols also detail the expression and purification of bacterial, viral, and eukaryotic DNA topoisomerases from a variety of sources, including baculovirus, and bacterial, yeast and mammalian expression systems. A companion volume, DNA Topoisomerase Protocols, v. 2: Enzymology and Drugs, provides detailed protocols to study the catalytic activities of DNA topoisomerases, as well as their specific interactions with topoisomerase-targeted antitumor and antibacterial drugs. DNA Topoisomerase Protocols, I: DNA Topology and Enzymes will be of immense value to the many basic scientists and clinicians who want better to understand and to exploit proficiently the wealth of recent discoveries about chromatin structure and its relation to gene expression, about DNA topoisomerases as the targets of antitumor and antibacterial agents, and about DNA repair. Beginning with the Escherichia coli co protein, or bacterial DNA topoisomerase I, an ever-increasing number of enzymes has been identified that catalyze changes in the linkage of DNA strands. DNA topoisomerases are ubiquitous in nature and have been shown to play critical roles in most p- cesses involving DNA, including DNA replication, transcription, and rec- bination. These enzymes further constitute the cellular targets of a number of clinically important antibacterial and anticancer agents. Thus, further studies of DNA topology and DNA topoisomerases are critical to advance our und- standing of the basic biological processes required for cell cycle progression, cell division, genomic stability, and development. In addition, these studies will continue to provide critical insights into the cytofoxic action of drugs that target DNA topoisomerases. Such mechanistic studies have already played an important role in the development and clinical application of antimicrobial and chemotherapeutic agents. The two volumes of DNA Topoisomerase Protocols are designed to help new and established researchers investigate all aspects of DNA topology and the function of these enzymes. The chapters are written by prominent investigators in the field and provide detailed background information and st- by-step experimental protocols. The topics covered in Volume I; DNA Topology and Enzymes, range from detailed methods to analyze various aspects of DNA structure, from linking number, knotting/unknotting, site-specific recombi- tion, and decatenation to the overexpression and purification of bacterial and eukaryotic DNA topoisomerases from a variety of cell systems and tissues.