title: Codon randomization for rapid exploration of chemical space in thiopeptide antibiotic variants.
abstract: Thiopeptide antibiotics exhibit a profound level of chemical diversity that is installed through cascades of posttranslational modifications on ribosomal peptides. Here, we present a technique to rapidly explore the chemical space of the thiopeptide GE37468 through codon randomization, yielding insights into thiopeptide maturation as well as structure and activity relationships. In this incarnation of the methodology, we randomized seven residues of the prepeptide-coding region, enabling the generation of 133 potential thiopeptide variants. Variant libraries were subsequently queried in two ways. First, high-throughput MALDI-TOF mass spectrometry was applied to colony-level expressions to sample mutants that permitted full maturation of the antibiotic. Second, the activity of producing mutants was detected in an antibiotic overlay assay. In total, 29 of the 133 variants produced mature compound, 12 of which retained antibiotic activity and 1 that had improved activity.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Armenise 608, Boston, MA 02115, USA.
authors.forename: Travis S
authors.initials: TS
authors.lastname: Young
authors.forename: Pieter C
authors.initials: PC
authors.lastname: Dorrestein
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Walsh
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D001483:Base Sequence; D003062:Codon / Q000235:genetics; D015870:Gene Expression; D005818:Genetic Engineering; D006801:Humans; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010456:Peptides, Cyclic / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology*; D013203:Staphylococcal Infections / Q000188:drug therapy; D013302:Streptomyces / Q000235:genetics; D013844:Thiazoles / Q000737:chemistry / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D003062:Codon; C096599:GE 37468 A; D010456:Peptides, Cyclic; D013844:Thiazoles