Cationic, Aromatic Acyl Donors for Directing Protein Acylation Using non-Covalent Interactions

Chemical acylation of biomolecules is one of the most important and fundamental bioconjugation reactions but generates statistical mixtures of conjugates that are difficult to control synthetically. Here, we explore the concept of small molecule acylation agent design in which the leaving group is capable of engaging in non-covalent interactions that can direct the acyl group to specific nucleophiles on a biomolecule. We achieved this through the systematic optimization of a cationic, aromatic ester that engages in peptide N-terminal labelling at physiological pH and can enable preparative scale peptide acylation in good yields. We explore the origin of the observed selectivity through the labelling of a model peptide through substituent effects using differing N-terminal residues. Expanding complexity to the acylation of proteins, protein mixtures, and a HELA cell proteome reveals that cationic, aromatic acyl donors label amine nucleophiles with very short reaction times and with highly distinct selectivity compared to classical N-hydroxysuccinimide (NHS) esters. Assessment across these mediums indicated that labelling selectivity/reactivity is likely directed by cation-π, dispersion, and electrostatic interactions, whilst NHS esters show sensitivity to steric environments.