Synthesis and analysis of stabilizing ligands for FKBP-derived destabilizing domains

• Published in: Bioorganic & medicinal chemistry Vol. 18; no. 2; pp. 759 - 761
• Main Authors: Grimley, Joshua S., Chen, Denise A., Banaszynski, Laura A., Wandless, Thomas J.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 15.01.2008; Elsevier
• Subjects: Animals; Biological and medical sciences; Biotechnology; Degradation; Fundamental and applied biological sciences. Psychology; Humans; Ligands; Methods. Procedures. Technologies; Mice; Protein engineering; Protein stability; Tacrolimus Binding Proteins - chemistry; Degradation; Protein engineering; Protein stability; Binding protein; Biological fixation; Ligand; Stabilization; Benzene derivatives; Ureas; Mutation; Morpholine derivatives; Chemical synthesis
• ISSN: 0960-894X; 0968-0896; 1464-3405; 1464-3391
• DOI: 10.1016/j.bmcl.2007.11.044

We recently engineered mutants of the FKBP12 protein that are rapidly degraded when expressed in cells. Shield-2 binds to destabilizing domains (DDs) and provides dose-dependent control of their expression levels. We recently identified mutants of the human FKBP12 protein that are unstable and rapidly degraded when expressed in mammalian cells. We call these FKBP mutants destabilizing domains (DDs), because their instability is conferred to any protein fused to the DDs. A cell-permeable ligand binds tightly to the DDs and prevents their degradation, thus providing small molecule control over intracellular protein levels. We now report the synthesis and functional characterization of a stabilizing ligand called Shield-2. The synthesis of Shield-2 is efficient, and this ligand binds to the FKBP(F36V) protein with a dissociation constant of 29 nM.