Elucidation of anthracyclinone biosynthesis by stepwise cloning of genes for anthracyclines from three different Streptomyces spp. The GenBank accession number for acmA reported in this paper is AF043550.

Kantola, Jaana; Kunnari, Tero; Hautala, Anne; Hakala, Juha; Ylihonko, Kristiina; Mäntsälä, Pekka

doi:10.1099/00221287-146-1-155

Cited by 38 publications

(47 citation statements)

References 32 publications

(19 reference statements)

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“…(Table 3) This compound, however, had 3 more sets of peaks in aliphatic proton region than SEK15 had, and the most characteristic peak was a doublet with 6 protons at δ = 0.8 ppm which corresponds to two terminal methyl groups in isobutyryl primed compound. Further spectroscopic analysis with 13 C NMR and COSY confirmed the structure of this compound as the expected SEK15-analog (YT243b, 4) along with the comparison with spectroscopic data for SEK15 34 .…”

Section: Biosynthesis Of Non-acetyl Primed Decaketide Backbones Usingmentioning

confidence: 79%

“…(Table 4) MS analysis of this compound was also consistent with its identity as an isobutyryl primed nonaketide. Further spectroscopic analysis of the compound via 13 C NMR, COSY, and HMBC confirmed its structure as shown in Scheme 2 (YT296a, 9, 2 mg/L). The 1 H and 13 C NMR spectra of the second compound suggested it was more closely related to aklanonic acid 38 , with additional peaks corresponding to the alkyl side chain, including a doublet corresponding to terminal methyl groups, analogous to YT243b (4) and YT292b (8).…”

Section: Biosynthesis Of Aklanonic Acid Analogsmentioning

confidence: 90%

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Engineered Biosynthesis of Aklanonic Acid Analogues

2005

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Aklanonic acid, an anthraquinone natural product, is a common advanced intermediate in the biosynthesis of several antitumor polyketide antibiotics, including doxorubicin and aclacinomycin A. Intensive semisynthetic and biosynthetic efforts have been directed toward developing improved analogs of these clinically important compounds. The primer unit of such polyfunctional aromatic polyketides is an attractive site for introducing novel chemical functionality, and attempts have been made to modify the primer unit by precursor directed biosynthesis or protein engineering of the polyketide synthase (PKS). We have previously demonstrated the feasibility of engineering bimodular aromatic PKSs capable of synthesizing unnatural hexaketides and octaketides. In this report we extend this ability by preparing analogs of aklanonic acid, a decaketide, and its methyl ester. For example, by recombining the R1128 initiation module with the dodecaketide-specific pradimicin PKS, the isobutyryl primed analog of aklanonic acid (YT296b, 10) and its methyl ester (YT299b, 12) were prepared. In contrast, elongation modules from dodecaketide-specific spore pigment PKSs were unable to interact with the R1128 initiation module. Thus, in addition to revealing a practical route to new anthracycline antibiotics, we also observed a fundamental incompatibility between antibiotic and spore pigment biosynthesis in the actinomycetes bacteria.

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Section: Biosynthesis Of Non-acetyl Primed Decaketide Backbones Usingmentioning

confidence: 79%

Section: Biosynthesis Of Aklanonic Acid Analogsmentioning

confidence: 90%

Engineered Biosynthesis of Aklanonic Acid Analogues

2005

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“…The role of snoaF was con®rmed by heterologous complementation of S. galilaeus H036 with the plasmid pSYE57 ( Table 2). Conversion of aklaviketone to aklavinone by SnoaF enabled the attachment of sugar moieties to the aglycone, and accumulation of aklavinone glycosides instead of aklaviketone, the normal product of H036 (Ylihonko et al 1994;Kantola et al 2000). 17.6(q) ± ± a The carbon numbering is depicted in Fig.…”

Section: Assignment Of Orfs To Functions In Aglycone Biosynthesismentioning

confidence: 99%

The entire nogalamycin biosynthetic gene cluster of Streptomyces nogalater: characterization of a 20-kb DNA region and generation of hybrid structures

et al. 2001

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Fragments spanning 20 kb of Streptomyces nogalater genomic DNA were characterized to elucidate the molecular genetic basis of the biosynthetic pathway of the anthracycline antibiotic nogalamycin. Structural analysis of the products obtained by expression of the fragments in S. galilaeus and S. peucetius mutants producing aclacinomycin and daunomycin metabolites, respectively, revealed hybrid compounds in which either the aglycone or the sugar moiety was modified. Subsequent sequence analysis revealed twenty ORFs involved in nogalamycin biosynthesis, of which eleven could be assigned to the deoxysugar pathway, four to aglycone biosynthesis, while the remaining five express products with unknown function. On the basis of sequence similarity and experimental data, the functions of the products of the newly discovered genes were determined. The results suggest that the entire biosynthetic gene cluster for nogalamycin is now known. Furthermore, the compounds obtained by heterologous expression of the genes show that it is possible to use the genes in combinatorial biosynthesis to create novel chemical structures for drug screening purposes.

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“…1) (19). Formation of the fourth, non-aromatic ring to yield alkaviketone during daunorubicin biosynthesis is catalyzed by the aklanonic acid methyl ester cyclase DnrD via distinct C-2/C-19 connectivity (20,21). The tetracenomycin family of compounds is constructed from fully aromatized intermediate tetracenomycin F1, but the regioselectivities are significantly different, starting with the first cyclization, which occurs between C-9 and C-14.…”

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confidence: 99%

Investigation of Early Tailoring Reactions in the Oxytetracycline Biosynthetic Pathway

Zhang

Watanabe

Wang

et al. 2007

Journal of Biological Chemistry

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Tetracyclines are aromatic polyketides biosynthesized by bacterial type II polyketide synthases. The amidated tetracycline backbone is biosynthesized by the minimal polyketide synthases and an amidotransferase homologue OxyD. Biosynthesis of the key intermediate 6-methylpretetramid requires two early tailoring steps, which are cyclization of the linearly fused tetracyclic scaffold and regioselective C-methylation of the aglycon. Using a heterologous host (CH999)/vector pair, we identified the minimum set of enzymes from the oxytetracycline biosynthetic pathway that is required to afford 6-methylpretetramid in vivo. Only two cyclases (OxyK and OxyN) are necessary to completely cyclize and aromatize the amidated tetracyclic aglycon. Formation of the last ring via C-1/C-18 aldol condensation does not require a dedicated fourth-ring cyclase, in contrast to the biosynthetic mechanism of other tetracyclic aromatic polyketides, such as daunorubicin and tetracenomycin. Acetylderived polyketides do not undergo spontaneous fourth-ring cyclization and form only anthracene carboxylic acids as demonstrated both in vivo and in vitro. OxyF was identified to be the C-6 C-methyltransferase that regioselectively methylates pretetramid to yield 6-methylpretetramid. Reconstitution of 6-methylpretetramid in a heterologous host sets the stage for a more systematic investigation of additional tetracycline downstream tailoring enzymes and is a key step toward the engineered biosynthesis of tetracycline analogs.Tetracyclines are aromatic polyketides biosynthesized by soil-borne Streptomyces bacteria (1, 2). It is well known that the carbon skeleton of an aromatic polyketide is assembled through stepwise decarboxylative condensation of malonate equivalents catalyzed by the minimal PKS, 2 which consists of a ketosynthase/chain length factor heterodimer (KS-CLF or KS ␣ -KS ␤ ), an acyl-carrier protein (ACP), and a malonyl-CoA:ACP acyltransferase (3). Dedicated tailoring enzymes transform the highly reactive poly-␤-ketone backbone into fused, richly substituted compounds (4). The biosynthesis of tetracyclines has been studied using blocked mutants of the chlorotetracycline producer Streptomyces aureofaciens (5-11). However, the underlying enzymology of several key tailoring steps that give rise to its structural features remain unresolved, including cyclization of the tetracyclic scaffold and C-methylation of C-6 to afford the key intermediate 6-methylpretetramid (Fig. 1). The oxytetracycline (oxy) biosynthetic gene cluster from Streptomyces rimosus has been completely sequenced, hence allowing a thorough investigation of the biochemical basis of these features (12-16).During tetracycline biosynthesis, the C-9 reduced decaketide backbone first undergoes C-7/C-12 intramolecular aldol condensation to fix the regioselectivity of the D ring followed by three sequential cyclization reactions to form a fully aromatized, tetracyclic intermediate pretetramid (Fig. 1). Biosynthesis of aureolic acids such as mithramycin presumably follows identical c...

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Elucidation of anthracyclinone biosynthesis by stepwise cloning of genes for anthracyclines from three different Streptomyces spp. The GenBank accession number for acmA reported in this paper is AF043550.

Cited by 38 publications

References 32 publications

Engineered Biosynthesis of Aklanonic Acid Analogues

Engineered Biosynthesis of Aklanonic Acid Analogues

The entire nogalamycin biosynthetic gene cluster of Streptomyces nogalater: characterization of a 20-kb DNA region and generation of hybrid structures

Investigation of Early Tailoring Reactions in the Oxytetracycline Biosynthetic Pathway

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