title: Towards Biocontained Cell Factories: An Evolutionarily Adapted Escherichia coli Strain Produces a New-to-nature Bioactive Lantibiotic Containing Thienopyrrole-Alanine.
abstract: Genetic code engineering that enables reassignment of genetic codons to non-canonical amino acids (ncAAs) is a powerful strategy for enhancing ribosomally synthesized peptides and proteins with functions not commonly found in Nature. Here we report the expression of a ribosomally synthesized and post-translationally modified peptide (RiPP), the 32-mer lantibiotic lichenicidin with a canonical tryptophan (Trp) residue replaced by the ncAA L-β-(thieno[3,2-b]pyrrolyl)alanine ([3,2]Tpa) which does not sustain cell growth in the culture. We have demonstrated that cellular toxicity of [3,2]Tpa for the production of the new-to-nature bioactive congener of lichenicidin in the host Escherichia coli can be alleviated by using an evolutionarily adapted host strain MT21 which not only tolerates [3,2]Tpa but also uses it as a proteome-wide synthetic building block. This work underscores the feasibility of the biocontainment concept and establishes a general framework for design and large scale production of RiPPs with evolutionarily adapted host strains.
authors.affiliation: Technische Universität Berlin, Institut für Chemie, Biologische Chemie, Straße des 17. Juni 124, Berlin, 10623, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Kuthning
authors.affiliation: Technische Universität Berlin, Institut für Chemie, Biokatalyse, Müller-Breslau-Straße 10, Berlin, 10623, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Durkin
authors.affiliation: Technische Universität Berlin, Institut für Chemie, Biokatalyse, Müller-Breslau-Straße 10, Berlin, 10623, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Oehm
authors.affiliation: Technische Universität Berlin, Institut für Chemie, Biokatalyse, Müller-Breslau-Straße 10, Berlin, 10623, Germany.
authors.forename: Michael G
authors.initials: MG
authors.lastname: Hoesl
authors.affiliation: Technische Universität Berlin, Institut für Chemie, Biokatalyse, Müller-Breslau-Straße 10, Berlin, 10623, Germany.
authors.forename: Nediljko
authors.initials: N
authors.lastname: Budisa
authors.affiliation: Technische Universität Berlin, Institut für Chemie, Biologische Chemie, Straße des 17. Juni 124, Berlin, 10623, Germany.
authors.forename: Roderich D
authors.initials: RD
authors.lastname: Süssmuth
mesh_terms: D000222:Adaptation, Physiological; D000409:Alanine / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000890:Anti-Infective Agents / Q000494:pharmacology; D001430:Bacteriocins / Q000494:pharmacology*; D005075:Biological Evolution; D004926:Escherichia coli / Q000187:drug effects / Q000502:physiology*; D008826:Microbial Sensitivity Tests; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011758:Pyrroles / Q000494:pharmacology*; D012270:Ribosomes / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D000890:Anti-Infective Agents; D001430:Bacteriocins; D010455:Peptides; D011758:Pyrroles; C521801:thienopyrrole; D000409:Alanine