Guengerich F Peter, Martin Martha V, Sohl Christal D, Cheng Qian
Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.
Nat Protoc. 2009;4(9):1245-51. doi: 10.1038/nprot.2009.121. Epub 2009 Aug 6.
Cytochrome P450 (P450) enzymes are important in the metabolism of steroids, vitamins, carcinogens, drugs and other compounds. Two of the commonly used assays in this field are the measurements of total P450 and NADPH-P450 reductase in biological preparations. A detailed protocol is presented for the measurement of P450 by its spectral properties, along with a protocol for measuring NADPH-P450 reductase by its NADPH-cytochrome c reduction activity. Each assay can be completed in 5-10 min. Detailed explanations for the rationale of particular sequences in the protocols are provided, along with potential confounding problems.
细胞色素P450(P450)酶在类固醇、维生素、致癌物、药物及其他化合物的代谢过程中发挥着重要作用。该领域常用的两种检测方法是测定生物制剂中的总P450和NADPH - P450还原酶。本文详细介绍了通过P450的光谱特性进行检测的方案,以及通过NADPH - 细胞色素c还原活性来检测NADPH - P450还原酶的方案。每项检测均可在5 - 10分钟内完成。文中还提供了对方案中特定步骤原理的详细解释以及可能存在的混淆问题。