Synergistic Copper-Organoboron Activation of Aspartic Acid Enabled Peptide Cyclization and Conjugation

Few advances have effectively addressed the challenge of selectively activating aspartic acid (Asp) side chains. The β-carboxyl group of Asp remains underutilized as a reactive handle due to its limited reactivity under biocompatible conditions, its susceptibility to competing aspartimide formation, and the difficulty of achieving site-selectivity among chemically indistinguishable carboxyl groups on glutamic acid (Glu) side chains and the C-terminus. Here, we report a copper/organoboron synergistic system that enables direct and selective activation of the Asp side chain within Asp-His motifs, forming Lys-Asp isopeptide bonds under neutral aqueous conditions and exhibiting broad compatibility with unprotected peptides containing all canonical amino acids, including Glu and Asp residues outside the recognition motif. The method is applied to the synthesis of cyclic peptides (14- to 38-membered rings), bicyclic architectures, and diverse bioactive sequences, including the RGD peptide. The utility of this Asp activation strategy extends to protein engineering, as demonstrated by the head-to-tail cyclization of the de novo-designed protein ABLE and protein-protein ligation mediated by a heterodimeric coiled-coil. Beyond cyclization, we identified a -DAGGDHLP- tag that exploits the activated Asp residue for site-specific peptide conjugation with small-molecule amines. This dual-reagent activation strategy provides an enzyme-free solution for Asp side-chain chemistry, expanding the synthetic toolkit for peptide and protein engineering.