The Biosynthesis of Teicoplanin-Type Glycopeptide Antibiotics: Assignment of P450 Mono-Oxygenases to Side Chain Cyclizations of Glycopeptide A47934

Abstract

Streptomyces toyocaensis produces A47934, a teicoplanin-like type-IV glycopeptide with antibiotic activity against methicillin-resistant Staphylococcus aureus. A47934 differs from the type-I vancomycin glycopeptides, which possess a tricyclic peptide backbone, by the presence of an additional ring closure between the aromatic amino acids 1 and 3. To elucidate the order of crosslinking reactions, P450 mono-oxygenase-inactivation mutants (DeltastaF, DeltastaG, DeltastaH, and DeltastaJ) of the A47934 producer were generated, and the accumulated intermediates were analyzed. Thus, the formation of each crosslink could unambiguously be assigned to a specific oxygenase. The structure of the released intermediates from the wild-type nonribosomal peptide synthetase assembly line facilitated the determination of the cyclization order. Unexpectedly, the additional ring closure in A47934, catalyzed by StaG, is the second oxygenase reaction.

Overproduction of Ristomycin A by Activation of a Silent Gene Cluster in Amycolatopsis japonicum MG417-CF17

“…These cross-linked glycopeptides contain three ether bridges (between amino acids 1 and 3, 2 and 4, and 4 and 6) and one C-C link (between amino acids 5 and 7). On the basis of the model proposed for the balhimycin and A47934 oxygenases (42,43) and using amino acid sequence similarity as a criterion, it is likely that OxyA is involved in the crosslinking of the aromatic residues of amino acids 2 and 4, while OxyB is involved in the cross-linking of amino acids 4 and 6 and OxyC is involved in the cross-linking of amino acids 5 and 7. The fourth encoded oxygenase (OxyE) catalyzes the type III and type IV glycopeptide-specific cross-linking between Hpg at position 1 and Dpg at position 3 (43) (Fig.…”

In vivo Mutational Analysis of the Mupirocin Gene Cluster Reveals Labile Points in the Biosynthetic Pathway: the “Leaky Hosepipe” Mechanism

“…MupC might thus be able to manifest activity on different substrates. Support for such a dual action of a tailoring enzyme comes from a recent study [30] on the P450 enzymes that control the formation of the biphenyl and diphenyl ether linkages in the biosynthesis of the vancomycin family of glycopeptide antibiotics. These studies have shown that the oxidative enzymes act in a defined sequence on specific substrates in wild-type (WT) organisms.…”

Unrivalled diversity: the many roles and reactions of bacterial cytochromes P450 in secondary metabolism

“…In these NRPS-biosynthesised heptapeptides, the three (vancomycin-type) or four (teicoplanin-type) crosslinks have been demonstrated to be inserted in vivo by cytochrome P450 enzymes, with each ring installed by a specic P450. [244][245][246][247][248] Furthermore, gene disruption experiments determined that a specic order of ring installation occurs during GPA biosynthesis (Fig. 21B).…”

“…Ring insertion commences with generation of the C-O-D ring by OxyB, with subsequent activity of the optional OxyE enzyme (installing the teicoplanin-type F-O-G ring) and the essential OxyA enzyme (installing the D-O-E ring), before terminating with insertion of the AB crosslink that is catalysed by OxyC. [244][245][246][247][248][249] These experiments also led to the hypothesis that the crosslinking performed by these P450 enzymes was likely to occur against NRPS bound (i.e. PCP-bound) peptides, which was conrmed for the OxyB enzyme from vancomycin biosynthesis by the work of Robinson and co-workers.…”