Photo-affinity labeling (PAL) in chemical proteomics: a handy tool to investigate protein-protein interactions (PPIs)

Abstract

Protein-protein interactions (PPIs) trigger a wide range of biological signaling pathways that are crucial for biomedical research and drug discovery. Various techniques have been used to study specific proteins, including affinity chromatography, activity-based probes, affinity-based probes and photo-affinity labeling (PAL). PAL has become one of the most powerful strategies to study PPIs. Traditional photocrosslinkers are used in PAL, including benzophenone, aryl azide, and diazirine. Upon photoirradiation, these photocrosslinkers (Pls) generate highly reactive species that react with adjacent molecules, resulting in a direct covalent modification. This review introduces recent examples of chemical proteomics study using PAL for PPIs.

Nonenzymatic synthesis of anomerically pure, mannosyl-based molecular probes for scramblase identification studies

“…As such, an attempt already showed great promise in a previous report by Rush et al from 2015 [10]. Similarly, we envisioned that these analogs could be used to capture MPD-recognizing proteins, including the scramblase, from a crude mixture of ER membrane proteins [11,12]. The captured proteins would be subsequently identified by mass spectrometry, and their function in MPD scrambling validated by biochemical and genetic approaches.…”

A bifunctional amino acid to study protein–protein interactions

“…Photo-cross-linking represents a powerful method to study PPIs, which converts the noncovalent interactions between a protein of interest and its binding partners into covalent linkages, facilitating the following identication and characterization processes. [4][5][6] Among all the frequently used photoreactive groups, diazirine is known for its small size and high reactivity. [7][8][9][10] These two features of diazirine ensure high crosslinking specicity and efficiency, as it can be installed close enough to the binding sites without inducing undesired steric hindrance, and thereby selectively and rapidly label the PPIs of interest upon photo-activation.…”

A versatile method for the UVA-induced cross-linking of acetophenone- or benzophenone-functionalized DNA

“…Photochemical cross-linking is a powerful strategy for studying biomolecule interactions as well as for anchoring a variety of ligands on different surfaces. A general photo-induced covalent cross-linking procedure can be achieved either by using an external photolinker 30 or by introducing a native photoactive functional group into a targeted molecule 31 , the latter being a more specific and efficient approach. Aryl ketones, aryl azides and diazirines are the most popular photoreactive groups used for the photo cross-linking 30 , 31 .…”

“…A general photo-induced covalent cross-linking procedure can be achieved either by using an external photolinker 30 or by introducing a native photoactive functional group into a targeted molecule 31 , the latter being a more specific and efficient approach. Aryl ketones, aryl azides and diazirines are the most popular photoreactive groups used for the photo cross-linking 30 , 31 . In fact, these have been used to study the cross-linking of DNA-DNA 32 or protein-DNA/RNA complexes 33 – 36 , to probe protein-protein interactions 37 , 38 or to attach a variety of biomolecules to polymer surfaces 39 , 40 .…”