Understanding thioamitide biosynthesis using pathway engineering and untargeted metabolomics

Abstract

Thiostreptamide S4 is a thioamitide, a family of promising antitumour ribosomally synthesised and post-translationally modified peptides (RiPPs). The thioamitides are one of the most structurally complex RiPP families, yet very...

Characterization of Histidine Functionalization and Its Timing in the Biosynthesis of Ribosomally Synthesized and Posttranslationally Modified Thioamitides

“…It is likely that TvaG methylates the unmodified His12 residue in 3 to dmHis in 2, followed by dmHis12 β-hydroxylation, which is catalyzed by TvaJ, to achieve the fully functionalized hdmHis in 1. Similar functional assignment was conducted independently by Koehnke et al 13 and Truman et al, 14 upon heterologous expression of a mutant tva cluster (in which the counterpart of tvaG or tvaJ was deleted) in a model Streptomyces strain. However, both TvaG and TvaJ need to be biochemically characterized.…”

“…14 While the exact timing remains to be biochemically determined, this PTM could be necessary for LP removal because the nonthioamidated analogues of 1, which are cyclized at the C-terminus and contain both AviCys and hdmHis residues, failed to be observed in the mutant Streptomyces strains that lacked thioamidation capability. 14 It is likely that thioamitide biosynthesis terminates with the activity of TvaK (Figure S29). For thioamitide maturation, this peptidase might not be sensitive to His functionalization because the precursors containing an unmodified His, dmHis, or hdmHis residue can be subjected to LP sequence removal for the production of 3, 2, or 1.…”

“…In contrast, prior peptide thioamidation appears to be unnecessary for His12 processing, as the reactions associated with TvaGJ activities can proceed with different substrates that are either thioamidated or nonthioamidated. Truman et al proposed that thioamidation of the precursor peptide serves as the first step according to gene deletion-based, detailed metabolomics analysis . While the exact timing remains to be biochemically determined, this PTM could be necessary for LP removal because the nonthioamidated analogues of 1 , which are cyclized at the C-terminus and contain both AviCys and hdmHis residues, failed to be observed in the mutant Streptomyces strains that lacked thioamidation capability .…”

“…Truman et al proposed that thioamidation of the precursor peptide serves as the first step according to gene deletion-based, detailed metabolomics analysis . While the exact timing remains to be biochemically determined, this PTM could be necessary for LP removal because the nonthioamidated analogues of 1 , which are cyclized at the C-terminus and contain both AviCys and hdmHis residues, failed to be observed in the mutant Streptomyces strains that lacked thioamidation capability . It is likely that thioamitide biosynthesis terminates with the activity of TvaK (Figure S29).…”

Beyond Soil-Dwelling Actinobacteria: Fantastic Antibiotics and Where to Find Them

“…No naturally occurring spliceotides have been isolated to date and there is no knowledge about their potential biological role, but coexpression in E. coli of these nif11 precursor peptides with their cognate radical SAM enzymes revealed their novel enzymatic activity, splicing the core peptide at one of the amide bonds and removing a tyramine equivalent to generate a β-amino acid in that position [294]. These results, alongside other genome mining studies revealing the presence of multiple RiPP cluster families, several of them still uncharacterized [295], indicate that there is plenty of cyanobacterial "chemical dark matter" still available to explore.…”