Department of Chemistry, Washington University in St. Louis, St. Louis, MO 63130, USA.
Department of Chemistry, Washington University in St. Louis, St. Louis, MO 63130, USA.
Methods. 2018 Jul 15;144:94-103. doi: 10.1016/j.ymeth.2018.05.016. Epub 2018 May 23.
Fast photochemical oxidation of proteins (FPOP) is a footprinting technique used in mass spectrometry-based structural proteomics. It has been applied to solve a variety of problems in different areas of biology. A FPOP platform requires a laser, optics, and sample flow path properly assembled to enable fast footprinting. Sample preparation, buffer conditions, and reagent concentrations are essential to obtain reasonable oxidations on proteins. FPOP samples can be analyzed by LC-MS methods to measure the modification extent, which is a function of the solvent-accessible surface area of the protein. The platform can be expanded to accommodate several new approaches, including dose-response studies, new footprinting reagents, and two-laser pump-probe experiments. Here, we briefly review FPOP applications and in a detailed manner describe the procedures to set up an FPOP protein footprinting platform.
快速光化学氧化蛋白质（FPOP）是一种基于质谱的结构蛋白质组学中的足迹技术。它已被应用于解决生物学不同领域的各种问题。FPOP 平台需要将激光、光学器件和样品流路正确组装，以实现快速足迹检测。样品制备、缓冲条件和试剂浓度对于获得蛋白质的合理氧化至关重要。FPOP 样品可以通过 LC-MS 方法进行分析，以测量修饰程度，这是蛋白质溶剂可及表面积的函数。该平台可以扩展以适应几种新方法，包括剂量反应研究、新的足迹试剂和双激光泵浦探测实验。在这里，我们简要回顾了 FPOP 的应用，并详细描述了建立 FPOP 蛋白质足迹检测平台的步骤。
