Non-equilibrium covalently crosslinked hydrogels are synthesized using carbodiimide fueled coupling of carboxylic acids to anhydrides which eventually dissipate by hydrolysis.
Transient changes in molecular geometry are key to the function of many important biochemical systems. Here, we show that diphenic acids undergo out-of-equilibrium changes in dihedral angle when reacted with a carbodiimide chemical fuel. Treatment of appropriately functionalized diphenic acids with EDC (N-(3-(dimethylamino)propyl)-N′-ethylcarbodiimide hydrochloride) yields the corresponding diphenic anhydrides, reducing the torsional angle about the biaryl bond by ∼45°, regardless of substitution. In the absence of steric resistance, the reaction is well-described by a simple mechanism; the resulting kinetic parameters can be used to derive important properties of the system, such as yields and lifetimes. The reaction tolerates steric hindrance ortho to the biaryl bond, although the competing formation of (transient) byproducts complicates quantitative analysis.
Targeted protein degradation (TPD) is a promising strategy for drug development. Most degraders function by forcing the association of the target protein (TP) with an E3 Ubiquitin ligase, which in favorable cases results in the poly-Ubiquitylation of the TP and its subsequent degradation by the 26S proteasome. Here we explore the feasibility of a different TPD strategy in which the TP is recruited directly to the proteasome without the requirement for poly-Ubiquitylation. Using an engineered cell line in which the HaloTag protein is fused to one of the Ubiquitin receptors, we show that native protein targets can be degraded in this fashion when the cells are exposed to a chemical dimerizer containing a chloroalkane and a TP ligand. The potential advantages and disadvantages of Ubiquitin-independent degraders vs. traditional proteolysis-targeting chimeras are discussed.
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