Elucidating the Mechanism of <i>cis</i> Double Bond Formation in Epothilone Biosynthesis

Tang, Li; Ward, Shannon L.; Chung, Loleta; Carney, John R.; Li, Yong; Reid, Ralph; Katz, Leonard

doi:10.1021/ja030503f

Cited by 73 publications

(54 citation statements)

References 7 publications

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“…Therefore, in order to predict the amino acids incorporated into the bacillaene structure, the nonribosomal code was extracted from the two bae adenylation domains (Table 4). For the first NRPS module of BaeJ, the analysis gave 100% identity with the glycine-specific code (31,46) while no similarity to known codes was found for the other BaeN module. These data give rise to the assumption that bacillaene should contain at least one glycine residue.…”

Section: Discoverymentioning

confidence: 86%

“…It is possible that these activities are provided by other proteins acting in trans. Such a complementation has also been suggested for other polyketides, such as chalcomycin (49) and epothilone (46). Thus, in order to explain the presence of a double bond in epothilone, Tang and coworkers proposed a mechanism consisting of module skipping, use of a DH domain of a downstream module, and subsequent iterative use of the same module for the next round of elongation (46).…”

Section: Vol 188 2006 Polyketide Synthesis In Bacillus Amyloliquefamentioning

confidence: 89%

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Structural and Functional Characterization of Three Polyketide Synthase Gene Clusters in Bacillus amyloliquefaciens FZB 42

et al. 2006

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Although bacterial polyketides are of considerable biomedical interest, the molecular biology of polyketide biosynthesis in Bacillus spp., one of the richest bacterial sources of bioactive natural products, remains largely unexplored. Here we assign for the first time complete polyketide synthase (PKS) gene clusters to Bacillus antibiotics. Three giant modular PKS systems of the trans-acyltransferase type were identified in Bacillus amyloliquefaciens FZB 42. One of them, pks1, is an ortholog of the pksX operon with a previously unknown function in the sequenced model strain Bacillus subtilis 168, while the pks2 and pks3 clusters are novel gene clusters. Cassette mutagenesis combined with advanced mass spectrometric techniques such as matrixassisted laser desorption ionization-time of flight mass spectrometry and liquid chromatography-electrospray ionization mass spectrometry revealed that the pks1 (bae) and pks3 (dif) gene clusters encode the biosynthesis of the polyene antibiotics bacillaene and difficidin or oxydifficidin, respectively. In addition, B. subtilis OKB105 (pheA sfp 0 ), a transformant of the B. subtilis 168 derivative JH642, was shown to produce bacillaene, demonstrating that the pksX gene cluster directs the synthesis of that polyketide.Environmental Bacillus amyloliquefaciens strain FZB 42 is distinguished from the domesticated model organism Bacillus subtilis 168 (23) by several features important for rhizosphere competence particularly by its abilities to suppress competitive organisms present in the plant rhizosphere (17, 21) and to promote plant growth (16). In a previous contribution (20), we have reported that B. amyloliquefaciens FZB 42 is a producer of three families of lipopeptides, surfactins, bacillomycins D, and fengycins, which are well-known secondary metabolites with mainly antifungal activity. They are also produced by numerous B. subtilis strains (48). Furthermore, three giant gene clusters containing genes with homology to polyketide synthase (PKS) genes of modular organization were identified but not assigned functional roles. Mutants of FZB 42 deficient in the synthesis of cyclic lipopeptides were unable to suppress phytopathogenic fungi but still retained their antibacterial potency.Polyketides belong to a large family of secondary metabolites that include many bioactive compounds with antibacterial, immunosuppressive, antitumor, or other physiologically relevant bioactivities. Their biosynthesis is accomplished by stepwise decarboxylative Claisen condensations between the extender unit and the growing polyketide chain, generating enzyme-bound ␤-ketoacyl intermediates. Before a subsequent round of chain extension, a variable set of modifying enzymes can locally introduce structural variety. Similar to the nonribosomal synthesis of peptides, the PKS multienzyme system uses acyl carrier proteins (ACPs) that are posttranslationally modified with the 4Ј-phosphopantetheine prosthetic group to channel the growing polyketide intermediate during elongation processes (3). Type I PKSs...

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Section: Discoverymentioning

confidence: 86%

Section: Vol 188 2006 Polyketide Synthesis In Bacillus Amyloliquefamentioning

confidence: 89%

Structural and Functional Characterization of Three Polyketide Synthase Gene Clusters in Bacillus amyloliquefaciens FZB 42

et al. 2006

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“…The cis double bond could arise from the Diels-Alder reaction or from the actions of enoyl-isomerase or reduction domains in modular PKS's during chain elongation [16][17][18]. The trans-to-cis isomerization can also be catalyzed by exogenous post PKS tailoring enzymes, as observed in phoslactomycin biosynthesis in Streptomyces, which is catalyzed by an enzyme with homology to a nicotinamide adenine dinucleotide (NAD)-dependent epimerase/dehydratase [19].…”

Section: Posmentioning

confidence: 99%

Functional Analysis of the Fusarielin Biosynthetic Gene Cluster

et al. 2016

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Abstract:Fusarielins are polyketides with a decalin core produced by various species of Aspergillus and Fusarium. Although the responsible gene cluster has been identified, the biosynthetic pathway remains to be elucidated. In the present study, members of the gene cluster were deleted individually in a Fusarium graminearum strain overexpressing the local transcription factor. The results suggest that a trans-acting enoyl reductase (FSL5) assists the polyketide synthase FSL1 in biosynthesis of a polyketide product, which is released by hydrolysis by a trans-acting thioesterase (FSL2). Deletion of the epimerase (FSL3) resulted in accumulation of an unstable compound, which could be the released product. A novel compound, named prefusarielin, accumulated in the deletion mutant of the cytochrome P450 monooxygenase FSL4. Unlike the known fusarielins from Fusarium, this compound does not contain oxygenized decalin rings, suggesting that FSL4 is responsible for the oxygenation.

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“…This direct Pks15/1-to-PpsA chain translocation would "skip" the ACP L domain of PpsA. Domain skipping has in fact been demonstrated in a few PKS systems (53)(54)(55). Moreover, this direct-capture pathway would have adaptive value because it would not require ATP for PHPA intermediate activation, an essential step in pathway A (step A2).…”

Section: Resultsmentioning

confidence: 99%

Biosynthesis of Cell Envelope-Associated Phenolic Glycolipids in Mycobacterium marinum

et al. 2015

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dPhenolic glycolipids (PGLs) are polyketide synthase-derived glycolipids unique to pathogenic mycobacteria. PGLs are found in several clinically relevant species, including various Mycobacterium tuberculosis strains, Mycobacterium leprae, and several nontuberculous mycobacterial pathogens, such as M. marinum. Multiple lines of investigation implicate PGLs in virulence, thus underscoring the relevance of a deep understanding of PGL biosynthesis. We report mutational and biochemical studies that interrogate the mechanism by which PGL biosynthetic intermediates (p-hydroxyphenylalkanoates) synthesized by the iterative polyketide synthase Pks15/1 are transferred to the noniterative polyketide synthase PpsA for acyl chain extension in M. marinum. Our findings support a model in which the transfer of the intermediates is dependent on a p-hydroxyphenylalkanoyl-AMP ligase (FadD29) acting as an intermediary between the iterative and the noniterative synthase systems. Our results also establish the p-hydroxyphenylalkanoate extension ability of PpsA, the first-acting enzyme of a multisubunit noniterative polyketide synthase system. Notably, this noniterative system is also loaded with fatty acids by a specific fatty acyl-AMP ligase (FadD26) for biosynthesis of phthiocerol dimycocerosates (PDIMs), which are nonglycosylated lipids structurally related to PGLs. To our knowledge, the partially overlapping PGL and PDIM biosynthetic pathways provide the first example of two distinct, pathwaydedicated acyl-AMP ligases loading the same type I polyketide synthase system with two alternate starter units to produce two structurally different families of metabolites. The studies reported here advance our understanding of the biosynthesis of an important group of mycobacterial glycolipids. Mycobacterial infections are responsible for devastating morbidity and mortality worldwide (1-5). A critical player in the ability of the mycobacteria to produce disease is a formidable cell envelope believed to be responsible for the intrinsic resilience of the mycobacteria to inhospitable environments, antimicrobial agents, and host immune defenses (6-12). Among the unique components found in the cell envelope of several pathogenic mycobacteria are two structurally related families of glycosylated and nonglycosylated lipids commonly referred to as phenolic glycolipids (PGLs) and phthiocerol dimycocerosates (PDIMs), respectively (for a review, see reference 13). PGLs and PDIMs have unusual lipid scaffolds consisting of ␤-diol-containing, long-chain, aliphatic polyketides esterified with long-chain, multimethylbranched fatty acids onto the diol functionality (Fig. 1). PGLs and PDIMs are found in several mycobacterial pathogens (e.g., Mycobacterium tuberculosis strains, Mycobacterium bovis, Mycobacterium leprae, and Mycobacterium marinum) and thought to be constituents of the characteristic mycobacterial outer membrane. Multiple lines of investigation have provided considerable support for the idea that PGLs and PDIMs are implicated in virulence via co...

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Elucidating the Mechanism of cis Double Bond Formation in Epothilone Biosynthesis

Cited by 73 publications

References 7 publications

Structural and Functional Characterization of Three Polyketide Synthase Gene Clusters in Bacillus amyloliquefaciens FZB 42

Structural and Functional Characterization of Three Polyketide Synthase Gene Clusters in Bacillus amyloliquefaciens FZB 42

Functional Analysis of the Fusarielin Biosynthetic Gene Cluster

Biosynthesis of Cell Envelope-Associated Phenolic Glycolipids in Mycobacterium marinum

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