Identification of a poly-cyclopropylglycine–containing peptide via bioinformatic mapping of radical S-adenosylmethionine enzymes

• Published in: The Journal of biological chemistry Vol. 298; no. 5; p. 101881
• Main Authors: Kostenko, Anastasiia, Lien, Yi, Mendauletova, Aigera, Ngendahimana, Thacien, Novitskiy, Ivan M., Eaton, Sandra S., Latham, John A.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.05.2022; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Sequence; bioinformatics; Carbon - metabolism; Computational Biology; Peptides - chemistry; Protein Processing, Post-Translational; radical-S-adenosylmethionine enzyme; ribosomally synthesized and posttranslationally modified peptide; S-Adenosylmethionine - metabolism; sequence similarity network; TVG biosynthesis; DMSO; EPR; MS; sequence similarity network; SSN; RiPP; radical-S-adenosylmethionine enzyme; EFI-GNT; ribosomally synthesized and posttranslationally modified peptide; TVG biosynthesis; bioinformatics; rSAM; EFI-EST; CPG; HSQC; BGC
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1016/j.jbc.2022.101881

Peptide-derived natural products are a large class of bioactive molecules that often contain chemically challenging modifications. In the biosynthesis of ribosomally synthesized and posttranslationally modified peptides (RiPPs), radical-SAM (rSAM) enzymes have been shown to catalyze the formation of ether, thioether, and carbon-carbon bonds on the precursor peptide. The installation of these bonds typically establishes the skeleton of the mature RiPP. To facilitate the search for unexplored rSAM-dependent RiPPs for the community, we employed a bioinformatic strategy to screen a subfamily of peptide-modifying rSAM enzymes which are known to bind up to three [4Fe-4S] clusters. A sequence similarity network was used to partition related families of rSAM enzymes into >250 clusters. Using representative sequences, genome neighborhood diagrams were generated using the Genome Neighborhood Tool. Manual inspection of bacterial genomes yielded numerous putative rSAM-dependent RiPP pathways with unique features. From this analysis, we identified and experimentally characterized the rSAM enzyme, TvgB, from the tvg gene cluster from Halomonas anticariensis. In the tvg gene cluster, the precursor peptide, TvgA, is comprised of a repeating TVGG motif. Structural characterization of the TvgB product revealed the repeated formation of cyclopropylglycine, where a new bond is formed between the γ-carbons on the precursor valine. This novel RiPP modification broadens the functional potential of rSAM enzymes and validates the proposed bioinformatic approach as a practical broad search tool for the discovery of new RiPP topologies.