Cytochrome P450 Protocols (Methods in Molecular Biology)

Ian Phillips and Elizabeth Shephard assemble a comprehensive collection of cutting-edge techniques and invaluable classic methods for the investigation of cytochromes P450. Described in detail by hands-on experimentalists for easy reproducibility, these methods include spectral analysis, purification and enzymatic assays, expression in heterologous systems, and the production and use of antibodies, as well as methods for quantification of gene expression, transfection of hepatocytes, and for the investigation of DNA-protein interactions and genetic polymorphism. In addition, because of the growing importance of in vitro systems in pharmacological toxicology, the book contains techniques for the culture of rodent and human hepatocytes and human epidermis. Cytochrome P450 induction as a biomarker for environmental pollution and the generation of mice with targeted gene disruptions complete this exhaustive collection of core techniques. Cytochrome P450 Protocols provides both novice and experienced researchers across many fields-toxicology, pharmacology, environmental biology, biochemistry, and molecular biology-all the tools needed to elucidate the crucial biological role played by the cytochromes P450 in the metabolism of therapeutic drugs, chemical carcinogens, and environmental pollutants.

Cytochromes P450 (CYPs) comprise a large superfamily of proteins that are of central importance in the detoxification or activation of a tremendous number of natural and synthetic hydrophobic xenobiotics, including many therapeutic drugs, chemical carcinogens and environmental pollutants. CYPs are important in mediating interactions between an organism and its chemical environment and in the regulation of physiological processes. Cytochrome P450 Protocols, Third Edition focuses on high-throughput methods for the simultaneous analysis of multiple CYPs, substrates or ligands. Although the emphasis is on CYPs of mammalian origin, it reflects an increasing interest in CYPs of bacterial species. Also included are chapters on cytochrome P450 reductase (the redox partner of CYPs) and the flavin-containing monooxygenases (FMOs), and metabolomic and lipidomic approaches for identification of endogenous substrates of CYPs (‘de-orphanizing’ CYP substrates). Written in the successful Methods in Molecular Biology™ series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls.

 

Authoritative and easily accessible, Cytochrome P450 Protocols, Third Edition provides a wide range of techniques accessible to researchers in fields as diverse as biochemistry, molecular biology, pharmacology, toxicology, environmental biology and genetics.