Evolutionary Spread of Distinct O‐methyltransferases Guides the Discovery of Unique Isoaspartate‐Containing Peptides, Pamtides

• Published in: Advanced science 2024-01, Vol.11 (2), p.e2305946-n/a
• Main Authors: Lee, Hyunbin, Park, Sho Hee, Kim, Jiyoon, Lee, Jaehak, Koh, Min Sun, Lee, Jung Ho, Kim, Seokhee
• Format: Article
• Language: English
• Subjects: Biosynthesis; Enzymes; genome mining; Genomes; graspetides; Isoaspartic Acid - genetics; Isoaspartic Acid - metabolism; Methyltransferases - genetics; Methyltransferases - metabolism; Molecular weight; pamtides; Peptides; Peptides - metabolism; protein L‐(iso)aspartyl O‐methyltransferase; Protein Processing, Post-Translational; Ribosomes - genetics; Ribosomes - metabolism; RiPP
• ISSN: 2198-3844; 2198-3844
• DOI: 10.1002/advs.202305946

Ribosomally synthesized and post‐translationally modified peptides (RiPPs) are a structurally diverse class of natural products with a distinct biosynthetic logic, the enzymatic modification of genetically encoded precursor peptides. Although their structural and biosynthetic diversity remains largely underexplored, the identification of novel subclasses with unique structural motifs and biosynthetic pathways is challenging. Here, it is reported that peptide/protein L‐aspartyl O‐methyltransferases (PAMTs) present in several RiPP subclasses are highly homologous. Importantly, it is discovered that the apparent evolutionary transmission of the PAMT gene to unrelated RiPP subclasses can serve as a basis to identify a novel RiPP subclass. Biochemical and structural analyses suggest that homologous PAMTs convert aspartate to isoaspartate via aspartyl‐O‐methyl ester and aspartimide intermediates, and often require cyclic or hairpin‐like structures for modification. By conducting homology‐based bioinformatic analysis of PAMTs, over 2,800 biosynthetic gene clusters (BGCs) are identified for known RiPP subclasses in which PAMTs install a secondary modification, and over 1,500 BGCs where PAMTs function as a primary modification enzyme, thereby defining a new RiPP subclass, named pamtides. The results suggest that the genome mining of proteins with secondary biosynthetic roles can be an effective strategy for discovering novel biosynthetic pathways of RiPPs through the principle of “guilt by association”. Comprehensive bioinformatic analysis demonstrates the close evolutionary relationship of peptide/protein L‐aspartyl O‐methyltransferases (PAMTs) found in biosynthetic gene clusters for ribosomally synthesized and post‐translationally modified peptides (RiPPs). Genome mining, heterologous expression, mass analysis, and multidimensional nuclear magnetic resonance analyses uncover pamtides, a unique subclass of RiPPs containing isoaspartyl moiety.