title: Site-directed and global incorporation of orthogonal and isostructural noncanonical amino acids into the ribosomal lasso peptide capistruin.
abstract: Expansion of the structural diversity of peptide antibiotics was performed through two different methods. Supplementation-based incorporation (SPI) and stop-codon suppression (SCS) approaches were used for co-translational incorporation of isostructural and orthogonal noncanonical amino acids (ncAAs) into the lasso peptide capistruin. Two ncAAs were employed for the SPI method and five for the SCS method; each of them probing the incorporation of ncAAs in strategic positions of the molecule. Evaluation of the assembly by HR-ESI-MS proved more successful for the SCS method. Bio-orthogonal chemistry was used for post-biosynthetic modification of capistruin congener Cap_Alk10 containing the ncAA Alk (Nε-Alloc-L-lysine) instead of Ala. A second-generation Hoveyda-Grubbs catalyst was used for an in vitro metathesis reaction with Cap_Alk10 and an allyl alcohol, which offers options for post-biosynthetic modifications. The use of synthetic biology allows for the in vivo production of new peptide-based antibiotics from an expanded amino acid repertoire.
authors.affiliation: Institut für Chemie, Technische Universität Berlin, 10623 Berlin (Germany).
authors.forename: Rashed S
authors.initials: RS
authors.lastname: Al Toma
authors.forename: Anja
authors.initials: A
authors.lastname: Kuthning
authors.forename: Matthias P
authors.initials: MP
authors.lastname: Exner
authors.forename: Alexander
authors.initials: A
authors.lastname: Denisiuk
authors.forename: Juliane
authors.initials: J
authors.lastname: Ziegler
authors.forename: Nediljko
authors.initials: N
authors.lastname: Budisa
authors.forename: Roderich D
authors.initials: RD
authors.lastname: Süssmuth
mesh_terms: D000409:Alanine / Q000737:chemistry; D019943:Amino Acid Substitution; D000596:Amino Acids / Q000737:chemistry*; D001671:Biochemistry / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000235:genetics; D008239:Lysine / Q000737:chemistry; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D000596:Amino Acids; D010455:Peptides; D011994:Recombinant Proteins; C533129:capistruin; D008239:Lysine; D000409:Alanine
keywords: amber stop-codon suppression; capistruin; lasso peptides; metathesis; noncanonical amino acids; peptides