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Polyketide Biosynthesis (polyketide + biosynthesis)
Selected AbstractsTotal Synthesis of Silyl-Protected Early Intermediates of Polyketide Biosynthesis,EUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 16 2010Karsten Krohn Abstract The ketal- or dithioketal-protected isocoumarins 15,18 gave the corresponding 1-naphthols 21,26 in their reactions with the acetoacetate (10) or pentane-2,4-dione (19) dianions and the acetone monoanion. Subjection of the dithioketal-protected ester 28 to Baker,Venkataraman reaction conditions led to the 8-deoxy tautomeric, protected forms 29/30 of the early decaketide antibiotic intermediate 2b. However, the dithioketal protecting groups could not be removed without destruction of the molecule. Consequently the silyl-protected unstable early tri- and tetracyclic decaketide biosynthesis intermediates 37a, 37b, and 38a (precursors of angucycline and anthracycline antitumor antibiotics) were prepared through silylation of 33a and 33b, to afford 34a and 34b, and subsequent treatment with acetylacetone dianion. The ultimate synthetic goal, the silyl-protected 2,3-dialkylated naphthol derivative 41, was achieved by selective elongation of the bottom chain of the bis-silyl-protected methyl ester 36 with acetylacetone dianion. [source] The Structural Basis for Docking in Modular Polyketide BiosynthesisCHEMBIOCHEM, Issue 3 2006Kira J. Weissman Dr. Abstract Polyketide natural products such as erythromycin and rapamycin are assembled on polyketide synthases (PKSs), which consist of modular sets of catalytic activities distributed across multiple protein subunits. Correct protein,protein interactions among the PKS subunits which are critical to the fidelity of biosynthesis are mediated in part by "docking domains" at the termini of the proteins. The NMR solution structure of a representative docking domain complex from the erythromycin PKS (DEBS) was recently solved, and on this basis it has been proposed that PKS docking is mediated by the formation of an intermolecular four - ,-helix bundle. Herein, we report the genetic engineering of such a docking domain complex by replacement of specific helical segments and analysis of triketide synthesis by mutant PKSs in vivo. The results of these helix swaps are fully consistent with the model and highlight residues in the docking domains that may be targeted to alter the efficiency or specificity of subunit,subunit docking in hybrid PKSs. [source] Analysis of intracellular short organic acid-coenzyme A esters from actinomycetes using liquid chromatography-electrospray ionization-mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 9 2007Je Won Park Abstract A method employing silicone oil density centrifugation, solid-phase extraction (SPE) cleanup, and LC-ESI-MS/MS analysis was developed for the rapid, selective, sensitive, and quantitative detection of an intracellular pool of short organic acid-CoA esters in actinomycetes. The detection limit was determined to be approximately 0.8 pmol (1.2 ng/ml) for each standard CoA-ester analyzed by the present LC-ESI-MS/MS method. A selected ion chromatogram for a typical fragment ion (m/z 428) specific to CoA-esters enabled the detection of eight intracellular CoA-esters involved in both primary and secondary metabolisms. The application of this method to bacterial metabolomic study is demonstrated by the profiling of the intracellular CoA-ester pools in the wild-type Streptomyces venezuelae strain producing polyketide antibiotics (methymycin and pikromycin), a polyketide synthase (PKS)-deleted S. venezuelae mutant, and a S. venezuelae mutant expressing the heterologous PKS genes. By quantifying the individual CoA-esterlevel in three different genotypes of the S. venezuela e strain, further insight could be gained into the role of CoA-estersin polyketide biosynthesis. This analytical approach can be extended to the quantification of the size and composition of in vivo CoA-ester pools in various microbes, and can provide a detailed understanding of the relationship between the in vivo CoA-ester pool and the production of pharmaceutically important polyketides. Copyright © 2007 John Wiley & Sons, Ltd. [source] Construction and performance of heterologous polyketide-producing K-12- and B-derived Escherichia coliLETTERS IN APPLIED MICROBIOLOGY, Issue 2 2010J. Wu Abstract Aims:,Escherichia coli has emerged as a viable heterologous host for the production of complex, polyketide natural compounds. In this study, polyketide biosynthesis was compared between different E. coli strains for the purpose of better understanding and improving heterologous production. Methods and Results:, Both B and K-12 E. coli strains were genetically modified to support heterologous polyketide biosynthesis [specifically, 6-deoxyerythronolide B (6dEB)]. Polyketide production was analysed using a helper plasmid designed to overcome rare codon usage within E. coli. Each strain was analysed for recombinant protein production, precursor consumption, by-product production, and 6dEB biosynthesis. Of the strains tested for biosynthesis, 6dEB production was greatest for E. coli B strains. When comparing biosynthetic improvements as a function of mRNA stability vs codon bias, increased 6dEB titres were observed when additional rare codon tRNA molecules were provided. Conclusions:,Escherichia coli B strains and the use of tRNA supplementation led to improved 6dEB polyketide titres. Significance and Impact of the Study:, Given the medicinal potential and growing field of polyketide heterologous biosynthesis, the current study provides insight into host-specific genetic backgrounds and gene expression parameters aiding polyketide production through E. coli. [source] Solution structure and proposed domain,domain recognition interface of an acyl carrier protein domain from a modular polyketide synthasePROTEIN SCIENCE, Issue 10 2007Viktor Y. Alekseyev Abstract Polyketides are a medicinally important class of natural products. The architecture of modular polyketide synthases (PKSs), composed of multiple covalently linked domains grouped into modules, provides an attractive framework for engineering novel polyketide-producing assemblies. However, impaired domain,domain interactions can compromise the efficiency of engineered polyketide biosynthesis. To facilitate the study of these domain,domain interactions, we have used nuclear magnetic resonance (NMR) spectroscopy to determine the first solution structure of an acyl carrier protein (ACP) domain from a modular PKS, 6-deoxyerythronolide B synthase (DEBS). The tertiary fold of this 10-kD domain is a three-helical bundle; an additional short helix in the second loop also contributes to the core helical packing. Superposition of residues 14,94 of the ensemble on the mean structure yields an average atomic RMSD of 0.64 ± 0.09 Å for the backbone atoms (1.21 ± 0.13 Å for all non-hydrogen atoms). The three major helices superimpose with a backbone RMSD of 0.48 ± 0.10 Å (0.99 ± 0.11 Å for non-hydrogen atoms). Based on this solution structure, homology models were constructed for five other DEBS ACP domains. Comparison of their steric and electrostatic surfaces at the putative interaction interface (centered on helix II) suggests a model for protein,protein recognition of ACP domains, consistent with the previously observed specificity. Site-directed mutagenesis experiments indicate that two of the identified residues influence the specificity of ACP recognition. [source] Platform biochemicals for a biorenewable chemical industryTHE PLANT JOURNAL, Issue 4 2008Basil J. Nikolau Summary The chemical industry is currently reliant on a historically inexpensive, petroleum-based carbon feedstock that generates a small collection of platform chemicals from which highly efficient chemical conversions lead to the manufacture of a large variety of chemical products. Recently, a number of factors have coalesced to provide the impetus to explore alternative renewable sources of carbon. Here we discuss the potential impact on the chemical industry of shifting from non-renewable carbon sources to renewable carbon sources. This change to the manufacture of chemicals from biological carbon sources will provide an opportunity for the biological research community to contribute fundamental knowledge concerning carbon metabolism and its regulation. We discuss whether fundamental biological research into metabolic processes at a holistic level, made possible by completed genome sequences and integrated with detailed structural understanding of biocatalysts, can change the chemical industry from being dependent on fossil-carbon feedstocks to using biorenewable feedstocks. We illustrate this potential by discussing the prospect of building a platform technology based upon a concept of combinatorial biosynthesis, which would explore the enzymological flexibilities of polyketide biosynthesis. [source] | |||||||||||||