title: An Orthogonal D2O-Based Induction System that Provides Insights into d-Amino Acid Pattern Formation by Radical S-Adenosylmethionine Peptide Epimerases.
abstract: Radical S-adenosyl methionine peptide epimerases (RSPEs) are an enzyme family that accomplishes regiospecific and irreversible introduction of multiple d-configured residues into ribosomally encoded peptides. Collectively, RSPEs can generate diverse epimerization patterns in a wide range of substrates. Previously, the lack of rapid methods to localize epimerized residues has impeded efforts to investigate the function and applicative potential of RSPEs. An efficient mass spectrometry-based assay is introduced that permits characterization of products generated in E. coli. Applying this to a range of non-natural peptide-epimerase combinations, it is shown that the d-amino acid pattern is largely but not exclusively dictated by the core peptide sequence, while the epimerization order is dependent on the enzyme-leader pair. RSPEs were found to be highly promiscuous, which allowed for modular introduction of peptide segments with defined patterns.
authors.affiliation: Institute of Microbiology, Eigenössische Technische Hochschule (ETH) Zurich, Vladimir-Prelog-Weg 1-5/10, 8093, Zurich, Switzerland.
authors.forename: Brandon I
authors.initials: BI
authors.lastname: Morinaka
authors.affiliation: Institute of Microbiology, Eigenössische Technische Hochschule (ETH) Zurich, Vladimir-Prelog-Weg 1-5/10, 8093, Zurich, Switzerland.
authors.forename: Marjan
authors.initials: M
authors.lastname: Verest
authors.affiliation: Institute of Microbiology, Eigenössische Technische Hochschule (ETH) Zurich, Vladimir-Prelog-Weg 1-5/10, 8093, Zurich, Switzerland.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Freeman
authors.affiliation: Insitut Pasteur, Collection des Cyanobactéries, Département de Microbiologie, 75015, Paris, France.
authors.forename: Muriel
authors.initials: M
authors.lastname: Gugger
authors.affiliation: Institute of Microbiology, Eigenössische Technische Hochschule (ETH) Zurich, Vladimir-Prelog-Weg 1-5/10, 8093, Zurich, Switzerland.
authors.forename: Jörn
authors.identifier: 0000-0002-2282-8154
authors.initials: J
authors.lastname: Piel
mesh_terms: D000596:Amino Acids / Q000096:biosynthesis* / Q000737:chemistry; D002238:Carbohydrate Epimerases / Q000737:chemistry / Q000378:metabolism*; D017666:Deuterium Oxide / Q000737:chemistry / Q000378:metabolism*; D005609:Free Radicals / Q000737:chemistry / Q000378:metabolism; D008968:Molecular Conformation; D012436:S-Adenosylmethionine / Q000737:chemistry / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D005609:Free Radicals; D012436:S-Adenosylmethionine; D002238:Carbohydrate Epimerases; D017666:Deuterium Oxide
keywords: epimerases; labeling; peptide biosynthesis; post-translation; radical-SAM