In Vivo Production of Diverse β-Amino Acid-Containing Proteins

The wide range of moieties installed in ribosomally synthesized and post-translationally modified peptides (RiPPs) suggests largely untapped potential for protein engineering. However, many RiPP maturases recognize target peptide precursors through an N-terminal leader sequence that is challenging to adapt to proteins. We have recently reported a family of enzymes that splice XYG sites in RiPPs to install alpha-keto-beta-amino acids. Backbone modifications influence diverse protein properties, yet the toolkit to install beta-amino acids is limited. Here we report their leader-independent incorporation into proteins in E. coli. Integrating an 11-residue splice tag into six different proteins permitted the site-selective introduction of beta-residues in vivo. The motif fusion at C-, N-terminal, and internal positions yielded various beta-residues. Our approach complements the few existing methods to introduce beta-amino acids or ketone-bearing moieties, suggesting diverse applications in chemical biology.

Automated summary

The study focuses on ribosomally synthesized and post-translationally modified peptides (RiPPs) and their untapped potential for protein engineering. The researchers report the incorporation of alpha-keto-beta-amino acids into proteins in E. coli using a family of enzymes that splice XYG sites in RiPPs. The approach offers new possibilities for introducing beta-amino acids or ketone-bearing moieties in chemical biology.