Electroaffinity Labeling: A New Platform for Chemoproteomic-based Target Identification

Abstract

Target identification is a critical pillar within the drug discovery process that involves deconvoluting the protein target of a pharmacologically active small molecule ligand. While photoaffinity labeling strategies have become the benchmark for target deconvolution of small molecules owing to their reliance on external activation to induce covalent protein capture, the process of target identification remains one of the most technically challenging aspects of early drug discovery. Thus, there is a strong demand for new technologies that allow for controlled activation of chemical probes to covalently label their protein target. Here, we introduce an electroaffinity labeling (ECAL) platform which leverages the use of a small, redox-active diazetidinone (DZE) functional group to enable chemoproteomic-based target identification of pharmacophores within live cell environments.

Design and Evaluation of a Cyclobutane Diazirine Alkyne Tag for Photoaffinity Labeling in Cells

“…Other approaches, such as using enantiomeric probes, can be used to increase the confidence in the specificity of these low-affinity interactions. Investigation into the reactivity profiles of exciting developments in other diazirine scaffolds, photoactivatable functional groups, and other activation chemistries will expand the scope of labeling experiments and assist in probe design in the future. Finally, probe design should take into consideration additional physicochemical properties, such as the lipophilicity and branching of the probe compound, the latter of which has been posited to increase the labeling specificity. , …”

Chemoproteomics, A Broad Avenue to Target Deconvolution

“…Recently, Baran and coworkers have developed an electroaffinity labeling platform that leverages the use of a small, redox-active diazetidinone functional group. [54] Diazetidinone can be electrochemically oxidized to covalently crosslink with its direct interacting partners in living cells, which opens up new avenues for capturing targets (Figure 4c).…”

“…The red-colored structure indicates the photoaffinity reactive group. c) The diazetidinone electrochemical label was developed by Y. Kawamata et al, [54] which can be electrochemically oxidized and produce reactive intermediate in biocompatible conditions. Chemoproteomics profiling of SR9009 was performed in such electroaffinity labelling (ECAL) way.…”

Photoaffinity Labeling Chemistries Used to Map Biomolecular Interactions

“…Modification of these functional groups has improved on their utility in PAL experiments. Recent innovations have opened new directions for labeling experiments that rely on triggered probe activation in cells, including blue light-activated, [83] IRactivated, [128] or electrochemically-activated chemistries, [129] each of which enable new experimental conditions that avoid damaging UV light or allow experiments in tissue samples that are normally inaccessible with traditional PAL chemistries. In addition, the development of more high-efficiency chemistries that target specific amino acids, such as 2,5-tetrazoles, can enable activity-based protein profiling experiments that are traditionally only performed with covalent labeling chemistries.…”