Repurposing Amide Bond-Forming Enzymes for Non-native Protein Modification

Site-specific protein modification techniques have led to breakthroughs across diverse fields, such as cell imaging, proteomics, biomaterials, and pharmaceutical protein and peptide syntheses. Complementing the versatility of chemical approaches, enzymatic strategies offer high regioselectivity, facilitating precise functionalization within intricate biological matrices. This review highlights recent progress in the use of amide bond-forming enzymes, particularly those that have been repurposed to expand the range of modifiable protein sites and modification types. Through enzyme-engineering methods including directed evolution, rational design, and computational redesign, alongside substrate processing involving acyl donors and acceptors, these enzymatic tools have been adapted to incorporate non-native protein targets and modifiers, thereby opening avenues in biotechnological applications.