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The tunicamycins are archetypal nucleoside antibiotics targeting bacterial peptidoglycan biosynthesis and eukaryotic protein N-glycosylation. Understanding the biosynthesis of their unusual carbon framework may lead to variants with improved selectivity. Here, we demonstrate in vitro recapitulation of key sugar-manipulating enzymes from this pathway. TunA is found to exhibit unusual regioselectivity in the reduction of a key α,β-unsaturated ketone. The product of this reaction is shown to be the preferred substrate for TunF--an epimerase that converts the glucose derivative to a galactose. In Streptomyces strains in which another gene (tunB) is deleted, the biosynthesis is shown to stall at this exo-glycal product. These investigations confirm the combined TunA/F activity and delineate the ordering of events in the metabolic pathway. This is the first time these surprising exo-glycal intermediates have been seen in biology. They suggest that construction of the aminodialdose core of tunicamycin exploits their enol ether motif in a mode of C-C bond formation not previously observed in nature, to create an 11-carbon chain.

Original publication

DOI

10.1038/nchem.1351

Type

Journal article

Journal

Nat Chem

Publication Date

20/05/2012

Volume

4

Pages

539 - 546

Keywords

Anti-Bacterial Agents, Bacterial Proteins, Biocatalysis, Carbohydrate Epimerases, Computational Biology, Hydro-Lyases, Ketones, Multigene Family, Protein Structure, Tertiary, Pseudomonas aeruginosa, Stereoisomerism, Substrate Specificity, Tunicamycin