Optical control of the ?2-adrenergic receptor with opto-prop-2: A cis-active azobenzene analog of propranolol

In this study, we synthesized and evaluated new photoswitchable ligands for the beta-adrenergic receptors ?1-AR and ?2-AR, applying an azologization strategy to the first-generation beta-blocker propranolol. The resulting compounds (Opto-prop-1, -2, -3) have good photochemical properties with high levels of light-induced trans-cis isomerization (>94%) and good thermal stability (t1/2 > 10 days) of the resulting cis-isomer in an aqueous buffer. Upon illumination with 360-nm light to PSScis, large differences in binding affinities were observed for photoswitchable compounds at ?1-AR as well as ?2-AR. Notably, Opto-prop-2 (VUF17062) showed one of the largest optical shifts in binding affinities at the ?2-AR (587-fold, cis-active), as recorded so far for photoswitches of G protein-coupled receptors. We finally show the broad utility of Opto-prop-2 as a light-dependent competitive antagonist of the ?2-AR as shown with a conformational ?2-AR sensor, by the recruitment of downstream effector proteins and functional modulation of isolated adult rat cardiomyocytes

Fragment Screening Yields a Small-Molecule Stabilizer of 14-3-3 Dimers That Modulates Client Protein Interactions

The development of protein-protein interaction (PPI) inhibitors has been a successful strategy in drug development. However, the identification of PPI stabilizers has proven much more challenging. Here we report a fragment-based drug screening approach using the regulatory hub-protein 14-3-3 as a platform for identifying PPI stabilizers. A homogenous time-resolved FRET assay was used to monitor stabilization of 14-3-3/peptide binding using the known interaction partner estrogen receptor alpha. Screening of an in-house fragment library identified fragment 2 (VUF15640) as a putative PPI stabilizer capable of cooperatively stabilizing 14-3-3 PPIs in a cooperative fashion with Fusicoccin-A. Mechanistically, fragment 2 appears to enhance 14-3-3 dimerization leading to increased client-protein binding. Functionally, fragment 2 enhanced potency of 14-3-3 in a cell-free system inhibiting the enzyme activity of the nitrate reductase. In conclusion, we identified a general PPI stabilizer targeting 14-3-3, which could be used as a tool compound for investigating 14-3-3 client protein interactions