Topoisomerases Methods and Protocols

This volume is intended to provide both fundamental methods for long-standing assays as well as recent advances in the field toward newer pathways. Chapters guide readers through methods tailored to studying prokaryotic topoisomerases including fundamental protocols for purification and basic characterization of prokaryotic enzymes, enzymology examining DNA cleavage, relaxation, decatenation, and binding to more complex assays dissecting cellular function and interactions, and bioinformatics protocol for examining protein sequence interdependencies with application across many different proteins beyond topoisomerases.

 Authoritative and cutting-edge, Topoisomerases: Methods and Protocols as a resource both to new investigators and to long-standing researchers in the field of topoisomerases.

Expression and One-Step Purification of His-tagged Bacterial Topoisomerase Subunits from Escherichia coli.- Bacterial RADAR Assay: Isolation and Detection of Intracellular Covalent Complexes.- Bacterial Topoisomerase I Growth Complementation Assay.- DNA Supercoiling Catalyzed by Bacterial Gyrase.- DNA Decatenation Catalyzed by Bacterial Topoisomerase IV.- DNA Cleavage Mediated by Bacterial Type II Topoisomerases.- Expression, purification, and measurement of DNA supercoil relaxation activity by eukaryotic topoisomerase II.- Plasmid DNA Cleavage Assay with Eukaryotic Topoisomerase II.- Topoisomerase II DNA Cleavage Assay Using Fluorescently Labeled Double-Stranded Oligonucleotides.- Plasmid DNA Binding Electrophoretic Mobility Shift Assay with Eukaryotic Topoisomerase II.- Topoisomerase II N-Terminal ATPase Clamp Stabilization.- Yeast Tools for Studying Type II Topoisomerases in Budding Yeast.- Use of Xenopus Egg Extracts to Study the Effects of Topoisomerase Poisons During Vertebrate DNA Replication.- Inducing and monitoring liquid-liquid phase separation by type II topoisomerases.- Analyzing Topoisomerase II Cleavage Complexes Using Flow Cytometry.- Kinetoplast DNA Decatenation Assay with Eukaryotic Topoisomerase II.- Recombinant Topoisomerase 2 Production Using Culture Human Cells.- Using PSICalc to Identify Protein Sequence Interdependencies.