Adapting the Glaser Reaction for Bioconjugation: Robust Access to Structurally Simple,Rigid Linkers

Copper‐mediated coupling between alkynes to generate a structurally rigid, linear 1,3‐diyne linkage has been known for over a century. However, the mechanistic requirement to simultaneously maintain Cu^I and an oxidant has limited its practical utility, especially for complex functional molecules in aqueous solution. We find that addition of a specific bpy‐diol ligand protects unprotected peptides from Cu^II‐mediated oxidative damage through the formation of an insoluble Cu^II gel which solves the critical challenge of applying Glaser coupling to substrates that are degraded by Cu^II. The generality of this method is illustrated through the conjugation of a series of polar and nonpolar labels onto a fully unprotected GLP‐1R agonist through a linear 7 Å diynyl linker.     

Shape-Shifting Patchy Particles

A facile method to synthesize shape-shifting patchy particles on the colloidal scale is described. The design is based on the solvent-induced shifting of the patch shape between concave and convex features. The initial concave patchy particles were synthesized in a water suspension by a swelling-induced buckling process. Upon exposure to different solvents, the patches were tuned reversibly to be either concave or convex. These particles can be assembled into chained, branched, zigzag, and cyclic colloidal superstructures in a highly site-specific manner by surface–liquid capillary bridging. The biphasic nature of the particles also enables site-selective surface functionalization.     

Enzyme‐Mediated,Site‐Specific Protein Coupling Strategies for Surface‐Based Binding Assays

Covalent surface immobilization of proteins for binding assays is typically performed non‐specifically via lysine residues. However, receptors that either have lysines near their binding pockets, or whose presence at the sensor surface is electrostatically disfavoured, can be hard to probe. To overcome these limitations and to improve the homogeneity of surface functionalization, we adapted and optimized three different enzymatic coupling strategies (4′‐phosphopantetheinyl transferase, sortase A, and asparaginyl endopeptidase) for biolayer interferometry surface modification. All of these enzymes can be used to site‐specifically and covalently ligate proteins of interest via short recognition sequences. The enzymes function under mild conditions and thus immobilization does not affect the receptors’ functionality. We successfully employed this enzymatic surface functionalization approach to study the binding kinetics of two different receptor–ligand pairs.