title: The thiostrepton A tryptophan methyltransferase TsrM catalyses a cob(II)alamin-dependent methyl transfer reaction.
abstract: Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a novel class of natural products including several antibiotics and bacterial toxins. In countless RiPP biosynthetic pathways, cobalamin-dependent radical SAM (B12/rSAM) enzymes play a pivotal role. In the biosynthetic pathway of the antibiotic and anti-cancer agent thiostrepton A, TsrM, a B12/rSAM enzyme, catalyses the transfer of a methyl group to an electrophilic carbon atom of tryptophan. Here we show that methylcob(III)alamin is the probable physiological enzyme cofactor, and cob(II)alamin rather than cob(I)alamin is a key reaction intermediate. Furthermore, we establish that TsrM and a triple-alanine mutant alkylate cob(II)alamin efficiently leading to the synthesis of MeCbl. Exploiting TsrM substrate ambiguity, we demonstrate that TsrM does not catalyse substrate H-atom abstraction like most radical SAM enzymes. Based on these data, we propose an unprecedented radical-based C-methylation mechanism, which further expands the chemical versatility of rSAM enzymes.
authors.affiliation: INRA, ChemSyBio, UMR 1319 Micalis, F-78350 Jouy-en-Josas, France.|AgroParisTech, ChemSyBio, UMR Micalis, F-78350 Jouy-en-Josas, France.
authors.forename: Alhosna
authors.initials: A
authors.lastname: Benjdia
authors.affiliation: INRA, ChemSyBio, UMR 1319 Micalis, F-78350 Jouy-en-Josas, France.|AgroParisTech, ChemSyBio, UMR Micalis, F-78350 Jouy-en-Josas, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Pierre
authors.affiliation: Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan 48109-0600, USA.
authors.forename: Carmen
authors.initials: C
authors.lastname: Gherasim
authors.affiliation: INRA, ChemSyBio, UMR 1319 Micalis, F-78350 Jouy-en-Josas, France.|AgroParisTech, ChemSyBio, UMR Micalis, F-78350 Jouy-en-Josas, France.
authors.forename: Alain
authors.initials: A
authors.lastname: Guillot
authors.affiliation: INRA, ChemSyBio, UMR 1319 Micalis, F-78350 Jouy-en-Josas, France.|AgroParisTech, ChemSyBio, UMR Micalis, F-78350 Jouy-en-Josas, France.
authors.forename: Manon
authors.initials: M
authors.lastname: Carmona
authors.affiliation: Metalloproteins Unit, Institut de Biologie Structurale, UMR5075, CEA, CNRS, Univ. Grenoble-Alpes. 71, Avenue des Martyrs, CS 10090, 38044 Grenoble, France.
authors.forename: Patricia
authors.initials: P
authors.lastname: Amara
authors.affiliation: Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan 48109-0600, USA.
authors.forename: Ruma
authors.initials: R
authors.lastname: Banerjee
authors.affiliation: INRA, ChemSyBio, UMR 1319 Micalis, F-78350 Jouy-en-Josas, France.|AgroParisTech, ChemSyBio, UMR Micalis, F-78350 Jouy-en-Josas, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Berteau
mesh_terms: D003067:Coenzymes; D004926:Escherichia coli; D008780:Methyltransferases / Q000378:metabolism*; D013883:Thiostrepton / Q000096:biosynthesis*; D014364:Tryptophan / Q000378:metabolism*; D014805:Vitamin B 12 / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D003067:Coenzymes; C097751:cob(II)alamin; D014364:Tryptophan; C019476:mecobalamin; D008780:Methyltransferases; D013883:Thiostrepton; D014805:Vitamin B 12