Skip to main content
. 2016 Oct 14;138(44):14609–14615. doi: 10.1021/jacs.6b06645

Figure 2.

Figure 2

Evaluating the efficiency and selectivity of photoaffinity-labeling of recombinant proteins by the small-molecule probes. (a) Evaluating the BTK labeling efficiency using in-gel fluorescence (top panels). For reaction setup, 0.5 μg of BTK (final concentration ≈0.1 μM), 0.2 μM small-molecule probe, 10 μM Dasatinib (for competition only) in 50 μL of PBS were used. For photoirradiation, a hand-held UV lamp with a wavelength of 302 nm for ACT (5 min) and BP (20 min) or 365 nm for DA (10 min) was used. (b) Evaluating the BRD4 labeling efficiency using in-gel fluorescence (top panels). For reaction setup, 0.4 μg of BRD4 (final concentration ≈0.4 μM), 0.2 μM small-molecule probe, 5 μg of BSA, and 10 μM JQ-1 (for competition only) in 50 μL of PBS were used. The equal loading of proteins was verified by SYPRO Ruby staining of the same gels (bottom panels). See Supporting Information for procedures of click chemistry with TAMRA-azide and polyacrylamide gel electrophoresis.