Selective N‐Terminal Modification of Peptides and Proteins Using Fatty Acyl Phosphates

Abstract The selective modification of proteins and peptides is an important chemical biology tool with a wide variety of applications, including the production of biopharmaceuticals or the study of post‐translational modifications. In particular, the selective acylation of the N‐terminus over side chains in peptides and proteins is a highly desirable but challenging reaction in this field. Current methods have a range of shortcomings, including lack of selectivity or narrow substrate scope. Here we report a biomimetic approach using the in situ enzymatic reagent activation (ERA) of carboxylic acids with ATP to generate acyl‐adenosine monophosphates. This method displays high selectivity for the N‐termini of peptides and proteins, including pharmaceutically relevant liraglutide, glucagon and insulin. The ERA acylation tolerates a broad range of unsubstituted and substituted fatty acids, including azido and dicarboxylic acids, thus making it suitable for N‐terminal bioorthogonal labelling strategies. Moreover, this strategy can also be applied to the modification of antibodies. In general, the ERA acylation is a versatile and bioorthogonal method that we envisage finding wider applications in the field of bioconjugation and the production of stable peptide and protein conjugates.