2026/01/16 by EuTchen Ang, Jun Shi Chang, Wei-En Huang +3 · 1 voice
Medicine · Biochemistry, Genetics and Molecular Biology · #Microbial Natural Products and Biosynthesis #RNA and protein synthesis mechanisms #Biochemical and Structural Characterization
paper · doi:10.1021/jacs.5c19988
openalex publication_date 2026/01/16 · openalex created_date 2026/01/17 · openalex updated_date 2026/07/04
Thiopeptides are a class of ribosomally synthesized and post-translationally modified peptides (RiPPs) that show promise for drug discovery. Their biosynthesis depends on leader peptide recognition, whereby multiple enzymes are recruited to process a core region into the mature natural product. Here, we identify sequence determinants of the leader peptide–enzyme dynamics in a thiopeptide biosynthesis. Using the flexible in vitro (FIT)-Laz translation platform, we examined how leader mutations influence modifications performed by the enzymes that together complete lactazole biosynthesis. Initial DNA template translations informed single amino acid saturation mutagenesis using mRNA display. This revealed leader mutations that modulate enzyme recognition, as validated by aligning enrichment scores with observed modification. This study provides important insights into the underexplored role of RiPP leader peptides and informs the design of improved pseudonatural product libraries.