title: Synthetic biotechnology to study and engineer ribosomal bottromycin biosynthesis.
abstract: Bottromycins represent a promising class of antibiotics binding to the therapeutically unexploited A-site of the bacterial ribosome. By inhibiting translation they are active against clinically important pathogens, such as vancomycin-resistant Enterococci. Structurally, bottromycins are heavily modified peptides exhibiting various unusual biosynthetic features. To set the stage for compound modification and yield optimization, we identified the biosynthetic gene cluster, used synthetic biotechnology approaches to establish and improve heterologous production, and generated analogs by pathway genetic engineering. We unambiguously identified three radical SAM methyltransferase-encoding genes required for various methylations at unactivated carbons yielding tert-butyl valine, methyl-proline, and β-methyl-phenylalanine residues, plus a gene involved in aspartate methyl-ester formation. Evidence for the formation of the exo-thiazole unit and for a macrocyclodehydration mechanism leading to amidine ring formation is provided.
authors.affiliation: Department of Microbial Natural Products, Helmholtz Institute for Pharmaceutical Research Saarland, Helmholtz Centre for Infection Research and Pharmaceutical Biotechnology, Saarland University, Saarbrücken, Saarland 66123, Germany.
authors.forename: Liujie
authors.initials: L
authors.lastname: Huo
authors.forename: Shwan
authors.initials: S
authors.lastname: Rachid
authors.forename: Marc
authors.initials: M
authors.lastname: Stadler
authors.forename: Silke C
authors.initials: SC
authors.lastname: Wenzel
authors.forename: Rolf
authors.initials: R
authors.lastname: Müller
mesh_terms: D000578:Amidines / Q000737:chemistry; D001483:Base Sequence; D001709:Biotechnology; D003500:Cyclization; D016983:Enterococcus / Q000235:genetics / Q000378:metabolism; D005818:Genetic Engineering; D008780:Methyltransferases / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D010456:Peptides, Cyclic / Q000096:biosynthesis; D012270:Ribosomes / Q000378:metabolism*; D012436:S-Adenosylmethionine / Q000378:metabolism; D013302:Streptomyces / Q000235:genetics / Q000378:metabolism
chemical_list: D000578:Amidines; D010456:Peptides, Cyclic; C024712:bottromycin; D012436:S-Adenosylmethionine; D008780:Methyltransferases