Vinylogous chain branching catalysed by a dedicated polyketide synthase module

• Published in: Nature (London) 2013-10, Vol.502 (7469), p.124-128
• Main Authors: Bretschneider, Tom, Heim, Joel B., Heine, Daniel, Winkler, Robert, Busch, Benjamin, Kusebauch, Björn, Stehle, Thilo, Zocher, Georg, Hertweck, Christian
• Format: Article
• Language: English
• Subjects: 631/45/173; 631/535/1266; 631/92/349; 631/92/60; Aquatic plants; Architectural engineering; Biosynthesis; Burkholderia - chemistry; Burkholderia - enzymology; Burkholderia - genetics; Carbon; Catalysis; Chemical properties; Crystallography, X-Ray; Enzymes; Fermentation; Humanities and Social Sciences; Lactones - metabolism; letter; Macrolides - chemistry; Mass spectrometry; Metabolites; Models, Molecular; multidisciplinary; Mutagenesis; Polyketide Synthases - genetics; Polyketide Synthases - metabolism; Polyketides; Protein research; Protein Structure, Tertiary; Proteins; Science; Seedlings; Structure; Synthesis; Toxins
• ISSN: 0028-0836; 1476-4687
• DOI: 10.1038/nature12588

This study shows the structural and biochemical characterization of a new type of polyketide synthase module that catalyses the vinylogous addition of a malonyl unit to an unsaturated thioester, generating a branch in the growing polyketide chain; this characterization provides a mechanism by which the structural diversity of polyketide natural products can be increased. A new route to polyketides Polyketide synthases (PKSs) are modular enzyme complexes that produce polyketides, a large class of secondary metabolites — otherwise known as natural products. In this manuscript, the authors identify and characterize a novel PKS module from the endofungal bacterium Burkholderia rhizoxinica that catalyses a vinylogous addition of a malonyl unit to an unsaturated thioester, generating a branch in the growing polyketide chain. This enzymological activity is very unusual; all previously studied PKS modules catalyse the head-to-tail fusion of acyl and malonyl units, yielding a linear polyketide chain. This newly discovered reaction provides a simple mechanism by which structural diversity of polyketide natural products can be increased. Bacteria use modular polyketide synthases (PKSs) to assemble complex polyketides, many of which are leads for the development of clinical drugs, in particular anti-infectives and anti-tumoral agents 1 . Because these multifarious compounds are notoriously difficult to synthesize, they are usually produced by microbial fermentation. During the past two decades, an impressive body of knowledge on modular PKSs 2 , 3 has been gathered that not only provides detailed insight into the biosynthetic pathways but also allows the rational engineering of enzymatic processing lines to yield structural analogues 4 , 5 . Notably, a hallmark of all PKS modules studied so far is the head-to-tail fusion of acyl and malonyl building blocks, which leads to linear backbones. Yet, structural diversity is limited by this uniform assembly mode. Here we demonstrate a new type of PKS module from the endofungal bacterium Burkholderia rhizoxinica that catalyses a Michael-type acetyl addition to generate a branch in the carbon chain. In vitro reconstitution of the entire PKS module, X-ray structures of a ketosynthase-branching didomain and mutagenesis experiments revealed a crucial role of the ketosynthase domain in branching the carbon chain. We present a trapped intermediary state in which acyl carrier protein and ketosynthase are covalently linked by the bra