Phage display and selection of lanthipeptides on the carboxy-terminus of the gene-3 minor coat protein

• Published in: Nature communications 2017-11, Vol.8 (1), p.1500-10, Article 1500
• Main Authors: Urban, Johannes H., Moosmeier, Markus A., Aumüller, Tobias, Thein, Marcus, Bosma, Tjibbe, Rink, Rick, Groth, Katharina, Zulley, Moritz, Siegers, Katja, Tissot, Kathrin, Moll, Gert N., Prassler, Josef
• Format: Article
• Language: English
• Subjects: 631/1647/2163; 631/45/611; 631/61/338/469; Amino acids; Antimicrobial agents; Bacteria; Bridges; C-Terminus; Coat protein; Cytoplasm; Drug development; E coli; Engineering; Enzymes; Genetic engineering; Gram-positive bacteria; Humanities and Social Sciences; Ligands; multidisciplinary; N-Terminus; Natural products; Peptides; Phage display; Phages; Post-translation; Proteins; Science; Science (multidisciplinary); Streptavidin; U-Plasminogen activator; Urokinase
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-017-01413-7

Ribosomally synthesized and post-translationally modified peptides (RiPPs) are an emerging class of natural products with drug-like properties. To fully exploit the potential of RiPPs as peptide drug candidates, tools for their systematic engineering are required. Here we report the engineering of lanthipeptides, a subclass of RiPPs characterized by multiple thioether cycles that are enzymatically introduced in a regio- and stereospecific manner, by phage display. This was achieved by heterologous co-expression of linear lanthipeptide precursors fused to the widely neglected C-terminus of the bacteriophage M13 minor coat protein pIII, rather than the conventionally used N-terminus, along with the modifying enzymes from distantly related bacteria. We observe that C-terminal precursor peptide fusions to pIII are enzymatically modified in the cytoplasm of the producing cell and subsequently displayed as mature cyclic peptides on the phage surface. Biopanning of large C-terminal display libraries readily identifies artificial lanthipeptide ligands specific to urokinase plasminogen activator (uPA) and streptavidin. Lanthipeptides are a class of cyclic post-translationally modified peptides with potential drug-like properties. Here the authors develop a phage display system by expressing lanthipeptide precursors as C-terminal fusions to the phage M13 coat protein pIII in E. coli along with the heterologous modifying enzymes.