Photoaffinity Labeling in Target- and Binding-Site Identification

Abstract

Photoaffinity labeling (PAL) using a chemical probe to covalently bind its target in response to activation by light has become a frequently used tool in drug discovery for identifying new drug targets and molecular interactions, and for probing the location and structure of binding sites. Methods to identify the specific target proteins of hit molecules from phenotypic screens are highly valuable in early drug discovery. In this review, we summarize the principles of PAL including probe design and experimental techniques for in vitro and live cell investigations. We emphasize the need to optimize and validate probes and highlight examples of the successful application of PAL across multiple disease areas.The use of photoaffinity labeling (PAL) in medicinal chemistry and drug discovery has recently come to fruition [1]. PAL is a powerful technique used for the study of proteinligand interactions, where it can identify unknown targets of ligands, assist in the elucidation of protein structures, functions and conformational changes as well as identify novel or alternative binding sites in proteins [2]. In the current review, we will discuss the general principles of photoaffinity labeling concerning photoaffinity probe design and experimental strategies and the performance of the different photoaffinity groups available. We then focus on examples of the successful application of PAL in relation to the identification of molecular targets of small molecules, discovery of off-target interactions and the classification and structural elucidation of binding sites. The majority of examples presented here were published within the past 15 years, and greater emphasis has been given to recent examples illustrative of the general approaches. Photoaffinity probe designPAL is the use of a chemical probe that can covalently bind to its target in response to activation by light [3]. This is made possible by the incorporation of a photoreactive group within an otherwise reversibly binding probe compound. On irradiation with a specific wavelength of light, the photogroup forms a reactive intermediate that rapidly reacts with and binds to the nearest molecule, which ideally will be the target protein. Frank

Discovery of stimulator binding to a conserved pocket in the heme domain of soluble guanylyl cyclase

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Tetrazole‐Based Probes for Integrated Phenotypic Screening, Affinity‐Based Proteome Profiling, and Sensitive Detection of a Cancer Biomarker

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