Enzymatic Peptide and Protein Bromination: The BromoTrp Tag

Abstract Bio‐orthogonal reactions for modification of proteins and unprotected peptides are of high value in chemical biology. The combination of enzymatic halogenation with transition metal‐catalyzed cross‐coupling provides a feasible approach for the modification of proteins and unprotected peptides. By a semirational protein engineering approach, variants of the tryptophan 6‐halogenase Thal were identified that enable efficient bromination of peptides with a C‐ terminal tryptophan residue. The substrate scope was explored using di‐, tri‐, and tetrapeptide arrays, leading to the identification of an optimized peptide tag we named BromoTrp tag. This tag was introduced into three model proteins. Preparative scale post‐translational bromination was possible with only a single cultivation and purification step using the bro minating E. coli coexpression system Brocoli . Palladium‐catalyzed Suzuki–Miyaura cross‐coupling of the bromoarene was achieved with Pd nanoparticle catalysts at 37 °C, highlighting the rich potential of this strategy for bio‐orthogonal functionalization and conjugation.