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Biosynthesis of Macrocyclic Peptides with C-Terminal β-Amino-α-keto Acid Groups by Three Different Metalloenzymes.

Dinh T NguyenLingyang ZhuDanielle L GrayToby J WoodsChandrashekhar PadhiKristen M FlattDouglas A MitchellWilfred A van der Donk
Published in: ACS central science (2024)
Advances in genome sequencing and bioinformatics methods have identified a myriad of biosynthetic gene clusters (BGCs) encoding uncharacterized molecules. By mining genomes for BGCs containing a prevalent peptide-binding domain used for the biosynthesis of ribosomally synthesized and post-translationally modified peptides (RiPPs), we uncovered a new compound class involving modifications installed by a cytochrome P450, a multinuclear iron-dependent non-heme oxidative enzyme (MNIO, formerly DUF692), a cobalamin- and radical S -adenosyl-l-methionine-dependent enzyme (B12-rSAM), and a methyltransferase. All enzymes were functionally expressed in Burkholderia sp. FERM BP-3421. Structural characterization demonstrated that the P450 enzyme catalyzed the formation of a biaryl C-C cross-link between two Tyr residues with the B12-rSAM generating β-methyltyrosine. The MNIO transformed a C-terminal Asp residue into aminopyruvic acid, while the methyltransferase acted on the β-carbon of this α-keto acid. Exciton-coupled circular dichroism spectroscopy and microcrystal electron diffraction (MicroED) were used to elucidate the stereochemical configuration of the atropisomer formed upon biaryl cross-linking. To the best of our knowledge, the MNIO featured in this pathway is the first to modify a residue other than Cys. This study underscores the utility of genome mining to isolate new macrocyclic RiPPs biosynthesized via previously undiscovered enzyme chemistry.
Keyphrases
  • amino acid
  • genome wide
  • healthcare
  • gene expression
  • copy number
  • single cell
  • cell wall
  • mass spectrometry
  • room temperature
  • single molecule
  • dna binding
  • silver nanoparticles
  • transcription factor
  • drug discovery