Vinylogous chain branching catalysed by a dedicated polyketide synthase module

Bretschneider, Tom; Heim, Joel B.; Heine, Daniel; Winkler, Robert; Busch, Benjamin; Kusebauch, Björn; Stehle, Thilo; Zocher, Georg; Hertweck, Christian

doi:10.1038/nature12588

Cited by 113 publications

(131 citation statements)

References 42 publications

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“…The rhizoxin AT-less type I PKS contains a β-branching KS with an appended B domain, which produces a β-branch lactone (SI Appendix, Fig. S1C) (12,13). In analogy, the glutarimide ring of iso-migrastatin (MGS) is installed by a β-branching KS performing similar β-branching chemistry (14).…”

Section: Significancementioning

confidence: 99%

Structural and evolutionary relationships of “AT-less” type I polyketide synthase ketosynthases

Lohman

Osipiuk

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Acyltransferase (AT)-less type I polyketide synthases (PKSs) break the type I PKS paradigm. They lack the integrated AT domains within their modules and instead use a discrete AT that acts in trans, whereas a type I PKS module minimally contains AT, acyl carrier protein (ACP), and ketosynthase (KS) domains. Structures of canonical type I PKS KS-AT didomains reveal structured linkers that connect the two domains. AT-less type I PKS KSs have remnants of these linkers, which have been hypothesized to be AT docking domains. Natural products produced by AT-less type I PKSs are very complex because of an increased representation of unique modifying domains. AT-less type I PKS KSs possess substrate specificity and fall into phylogenetic clades that correlate with their substrates, whereas canonical type I PKS KSs are monophyletic. We have solved crystal structures of seven AT-less type I PKS KS domains that represent various sequence clusters, revealing insight into the large structural and subtle amino acid residue differences that lead to unique active site topologies and substrate specificities. One set of structures represents a larger group of KS domains from both canonical and AT-less type I PKSs that accept amino acid-containing substrates. One structure has a partial AT-domain, revealing the structural consequences of a type I PKS KS evolving into an AT-less type I PKS KS. These structures highlight the structural diversity within the AT-less type I PKS KS family, and most important, provide a unique opportunity to study the molecular evolution of substrate specificity within the type I PKSs.biosynthesis | secondary metabolism | iso-migrastatin | leinamycin | oxazolomycin

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Section: Significancementioning

confidence: 99%

Structural and evolutionary relationships of “AT-less” type I polyketide synthase ketosynthases

Lohman

Osipiuk

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…57). Nonetheless, the structures of all three of the core domains have now been reported: the KS domain (one from the rhizoxin pathway solved as a didomain with a downstream branching (B) domain 58 and another monodomain KS fragment from the bacillaene PKS 59 ), a discrete AT domain 57 and multiple ACP domains 60,61 . In addition, structures of domains for several unusual tailoring activities from the bacillaene assembly line (a KR that reduces both α-and β-keto groups 62 and an enoyl-isomerase (EI) that catalyzes a double-bond shift 63 ) have been reported.…”

Section: The Dissection Approach To Studying Functional Domainsmentioning

confidence: 97%

The structural biology of biosynthetic megaenzymes

Weissman¹

2015

Nat Chem Biol

185

165

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The modular polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs) are among the largest and most complicated enzymes in nature. In these biosynthetic systems, independently folding protein domains, which are organized into units called 'modules', operate in assembly-line fashion to construct polymeric chains and tailor their functionalities. Products of PKSs and NRPSs include a number of blockbuster medicines, and this has motivated researchers to understand how they operate so that they can be modified by genetic engineering. Beginning in the 1990s, structural biology has provided a number of key insights. The emerging picture is one of remarkable dynamics and conformational programming in which the chemical states of individual catalytic domains are communicated to the others, configuring the modules for the next stage in the biosynthesis. This unexpected level of complexity most likely accounts for the low success rate of empirical genetic engineering experiments and suggests ways forward for productive megaenzyme synthetic biology.

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“…Recently, β-additions, with carbon nucleophiles (i.e., Michael addition), to generate an alkyl branch in the growing polyketide chain have been observed (40)(41)(42). On the basis of the absolutely conserved KS-ACP-DUF-SH-TE domain architecture of the PKS module-8 of LnmJ and its homologs (Figs.…”

Section: Discussionmentioning

confidence: 99%

C-S bond cleavage by a polyketide synthase domain

Lohman

Liu

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Leinamycin (LNM) is a sulfur-containing antitumor antibiotic featuring an unusual 1,3-dioxo-1,2-dithiolane moiety that is spiro-fused to a thiazole-containing 18-membered lactam ring. The 1,3-dioxo-1,2-dithiolane moiety is essential for LNM's antitumor activity, by virtue of its ability to generate an episulfonium ion intermediate capable of alkylating DNA. We have previously cloned and sequenced the lnm gene cluster from Streptomyces atroolivaceus S-140. In vivo and in vitro characterizations of the LNM biosynthetic machinery have since established that: (i) the 18-membered macrolactam backbone is synthesized by LnmP, LnmQ, LnmJ, LnmI, and LnmG, (ii) the alkyl branch at C-3 of LNM is installed by LnmK, LnmL, LnmM, and LnmF, and (iii) leinamycin E1 (LNM E1), bearing a thiol moiety at C-3, is the nascent product of the LNM hybrid nonribosomal peptide synthetase (NRPS)-acyltransferase (AT)-less type I polyketide synthase (PKS). Sulfur incorporation at C-3 of LNM E1, however, has not been addressed. Here we report that: (i) the bioinformatics analysis reveals a pyridoxal phosphate (PLP)-dependent domain, we termed cysteine lyase (SH) domain (LnmJ-SH), within PKS module-8 of LnmJ; (ii) the LnmJ-SH domain catalyzes C-S bond cleavage by using L-cysteine and L-cysteine S-modified analogs as substrates through a PLP-dependent β-elimination reaction, establishing L-cysteine as the origin of sulfur at C-3 of LNM; and (iii) the LnmJ-SH domain, sharing no sequence homology with any other enzymes catalyzing C-S bond cleavage, represents a new family of PKS domains that expands the chemistry and enzymology of PKSs and might be exploited to incorporate sulfur into polyketide natural products by PKS engineering.cysteine metabolism | enzyme mechanism | genome mining | pathway engineering | sulfur metabolism S ulfur is found in many primary metabolites, such as thiamin, biotin, molybdenum cofactors, lipoic acid, iron-sulfur clusters, and nucleosides including 2-thiocytidine, 4-thiouridine, and 2-methylthio-N 6 -isopentenyl adenosine (1). There is a wealth of information on how sulfur is incorporated into these primary metabolites, the key step of which is catalyzed by a desulfurase to produce a protein-bound cysteine persulfide by using cysteine as a substrate (2-5). Sulfur is also found in many secondary metabolites (interchangeably referred to as natural products here). The major groups of sulfur-containing natural products are peptides produced by ribosome or nonribosomal peptide synthetases (NRPSs). Sulfur incorporation into these natural products from intact cysteine or methionine is well characterized. However, sulfur incorporation into other natural products remains poorly understood. Only a few cases featured by C-S bond formation or C-S bond cleavage steps were reported to date, and most of the mechanisms require further investigation (6-15).Leinamycin (LNM) is a sulfur-containing antitumor antibiotic produced by Streptomyces atroolivaceus S-140 (16). The structure of LNM is characterized by an unusual 1,3-dioxo-1,2-dith...

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Vinylogous chain branching catalysed by a dedicated polyketide synthase module

Cited by 113 publications

References 42 publications

Structural and evolutionary relationships of “AT-less” type I polyketide synthase ketosynthases

Structural and evolutionary relationships of “AT-less” type I polyketide synthase ketosynthases

The structural biology of biosynthetic megaenzymes

C-S bond cleavage by a polyketide synthase domain

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