Trifunctional Label Transfer

Figure 28.10 Trifunctional label transfer agents contain two arms with terminal reactive groups and a third arm with a label or affinity tag, such as a biotin group. One of the reactive groups typically is thermoreactive to couple with bait proteins, while the second reactive group usually is photoreactive. The thermoreactive arm has a cleavable cross-bridge to facilitate release of the captured protein and transfer of the label of affinity tag to it.

Figure 28.14 A trifunctional label transfer reagent containing a thiol-reactive pyridyl disulfide group, a photo-reactive phenyl azide, and a biotin affinity tag. This compound can be used to label bait proteins through available thiol groups and capture interacting prey proteins by photoreactive conjugation.

Figure 6.2 The trifunctional reagent sulfo-SBED reacts with amine-containing bait proteins via its NHS ester side chain. Subsequent interaction with a protein sample and exposure to UV light can cause crosslink formation with a second interacting protein. The biotin portion provides purification or labeling capability using avidin or streptavidin reagents. The disulfide bond on the NHS ester arm provides cleavability using disulfide reductants, which effectively transfers the biotin label to an unknown interacting protein.

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