title: A transglutaminase homologue as a condensation catalyst in antibiotic assembly lines.
abstract: The unrelenting emergence of antibiotic-resistant bacterial pathogens demands the investigation of antibiotics with new modes of action. The pseudopeptide antibiotic andrimid is a nanomolar inhibitor of the bacterial acetyl-CoA carboxylase that catalyses the first committed step in prokaryotic fatty acid biosynthesis. Recently, the andrimid (adm) biosynthetic gene cluster was isolated and heterologously expressed in Escherichia coli. This establishes a heterologous biological host in which to rapidly probe features of andrimid formation and to use biosynthetic engineering to make unnatural variants of this important and promising new class of antibiotics. Bioinformatic analysis of the adm cluster revealed a dissociated biosynthetic assembly system lacking canonical amide synthases between the first three carrier protein domains. Here we report that AdmF, a transglutaminase (TGase) homologue, catalyses the formation of the first amide bond, an N-acyl-beta-peptide link, in andrimid biosynthesis. Hence, AdmF is a newly discovered biosynthetic enzyme that acts as a stand-alone amide synthase between protein-bound, thiotemplated substrates in an antibiotic enzymatic assembly line. TGases (enzyme class (EC) 2.3.2.13) normally catalyse the cross-linking of (poly)peptides by creating isopeptidic bonds between the gamma-carboxamide group of a glutamine side chain of one protein and various amine donors, including lysine side chains. To the best of our knowledge, the present study constitutes the first report of a TGase-like enzyme recruited for the assembly of an antibiotic. Moreover, genome mining using the AdmF sequence yielded additional TGases in unassigned natural product biosynthetic pathways. With many more microbial genomes being sequenced, such a strategy could potentially unearth biosynthetic pathways producing new classes of antibiotics.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: Pascal D
authors.initials: PD
authors.lastname: Fortin
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Walsh
authors.forename: Nathan A
authors.initials: NA
authors.lastname: Magarvey
mesh_terms: D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000737:chemistry / Q000145:classification; D001419:Bacteria / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D001688:Biological Products / Q000096:biosynthesis / Q000737:chemistry; D002384:Catalysis; D004926:Escherichia coli; D005798:Genes, Bacterial / Q000235:genetics; D005810:Multigene Family / Q000235:genetics; D010649:Phenylalanine / Q000737:chemistry / Q000378:metabolism; D011090:Polyenes / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011758:Pyrroles / Q000737:chemistry / Q000378:metabolism; D011503:Transglutaminases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D001688:Biological Products; D011090:Polyenes; D011758:Pyrroles; C088069:andrimid; D010649:Phenylalanine; D011503:Transglutaminases