Mapping Ligand–receptor Interfaces: Approaching the Resolution Limit of Benzophenone‐based Photoaffinity Scanning

Abstract
Photoaffinity crosslinking has yielded important insights in the study of G protein‐coupled receptors and the mode of ligand binding. The most widely used photolabile moiety is p‐benzoylphenylalanine largely because of its reportedly high site specificity, reduced reactivity to water and light, photokinetics, and ease of incorporation into peptide ligands during synthesis. However, in the course of our studies directed at characterizing the binding of parathyroid hormone to its cognate G protein‐coupled receptor, we find that inherent properties of p‐benzoylphenylalanine, such as its size and conformational flexibility, limit the resulting resolution of the ligand–receptor structure. Here, we examine and define these limits.