Identification and Utility of FdmR1 as a
            <i>Streptomyces</i>
            Antibiotic Regulatory Protein Activator for Fredericamycin Production in
            <i>Streptomyces griseus</i>
            ATCC 49344 and Heterologous Hosts

Chen, Yihua; Wendt-Pienkowski, Evelyn; Shen, Ben

doi:10.1128/jb.00592-08

Cited by 83 publications

(73 citation statements)

References 46 publications

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“…Inactivation of FdmR1 in the fdm cluster abolished fredericamycin biosynthesis, supporting its function as the transcriptional activator. 48 The expression and modification of PnxR2 was apparently necessary for the initiation of transcription of the pnx cluster.…”

Section: Regulators Transporters and Othersmentioning

confidence: 99%

Cloning of the biosynthetic gene cluster for naphthoxanthene antibiotic FD-594 from Streptomyces sp. TA-0256

et al. 2010

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FD-594 is an unique pyrano [4¢,3¢:6,7]naphtho [1,2-b]xanthene polyketide with a trisaccharide of 2,6-dideoxysugars. In this study, we cloned the FD-594 biosynthetic gene cluster from the producer strain Streptomyces sp. TA-0256 to investigate its biosynthesis. The identified pnx gene cluster was 38 143 bp, consisting of 40 open reading frames, including a minimal PKS gene, TDP-olivose biosynthetic genes, two glycosyltransferase genes, two methyltransferase genes and many oxygenase/reductase genes. Most of these enzymes coded in the pnx cluster were reasonably assigned to a plausible biosynthetic pathway for FD-594, in which an unique ring opening process via Baeyer-Villiger-type oxidation catalyzed by a putative flavin adenine dinucleotide (FAD)-dependent monooxygenase, is speculated to lead to the unique xanthene structure. To clarify the involvement of pnx genes in the FD-594 biosynthesis, a glycosyltransferase, PnxGT2, and a methyltransferase, PnxMT2, were characterized enzymatically with the recombinant proteins expressed in Escherichia coli. As a result, PnxGT2 catalyzed the triple olivose transfers to the FD-594 aglycon with TDP-olivose as the glycosyl donor to afford triolivoside. Surprisingly, in the PnxGT2 enzymatic reaction, tetraolivoside and pentaolivoside were significantly detected along with the expected triolivoside. To our knowledge, PnxGT2 is the first contiguous oligosaccharide-forming glycosyltransferase in secondary metabolism. Furthermore, addition of PnxMT2 and S-adenosyl-L-methionine into the PnxGT2 reaction mixture afforded natural FD-594 to confirm that the PnxGT2 reaction product was the expected regiospecifically glycosylated compound. Consequently, the identified pnx gene cluster appears to be involved in FD-594 biosynthesis.

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Section: Regulators Transporters and Othersmentioning

confidence: 99%

Cloning of the biosynthetic gene cluster for naphthoxanthene antibiotic FD-594 from Streptomyces sp. TA-0256

et al. 2010

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“…S1). Finally, the previously reported plasmid pBS4045, which overproduces the fdm activator FdmR1 (25), was introduced into SB4027 via protoplast transformation, affording the ⌬fdmV variant SB4028.…”

Section: Methodsmentioning

confidence: 99%

“…Plasmids pGEM-3Zf(ϩ) (Promega), pSPORT1 (Invitrogen), and pET29a(ϩ) (Novagen) were from commercial sources. Plasmids pBS4028 (17), pBS4045 (25), pSET151 (23), pSET152 (23), pUWL201PW (26), and pWHM1250 (27) were described previously. Ampicillin at 100 g/ml, apramycin at 50 g/ml, kanamycin at 50 g/ml, and thiostrepton at 10 g/ml were used, respectively, in liquid media for selection (21,23).…”

Section: Methodsmentioning

confidence: 99%

Characterization of FdmV as an Amide Synthetase for Fredericamycin A Biosynthesis in Streptomyces griseus ATCC 43944

Chen

Wendt-Pienkowski

et al. 2010

Journal of Biological Chemistry

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Fredericamycin (FDM)AAmide bonds are common in natural products and represent fundamental linkages for numerous ribosomal and nonribosomal peptide-derived molecules; they are formed by the ribosome or by the condensation domain of nonribosomal peptide synthetases, respectively. Amide bonds can also be introduced into diverse natural products via different manifolds (Fig. 1). For instance, AdmF, a transglutaminase homolog, catalyzes amide bond formation in andrimid, highlighting a novel condensation strategy (1, 2). In the ansamycin-type antibiotics, the macrolactam ring in rifamycin is formed by an amide synthetase, RifF (3); whereas in some other macrolactams, like vicenistatin, the amide is proposed to result from thioesterase domain docking at the C terminus of the polyketide synthase (4). Alternative scenarios invoke amide bond formation as a tailoring step following construction of the core scaffold. For instance, in the aminocoumarin antibiotics, the amide bonds linking the 3-amino-4,7-dihydroxycoumarin moiety and related acyl moieties are formed by amide synthetases such as NovL (for novobiocin) and its homologs CloL, CouL, and SimL (5, 6). Novel novobiocin analogs can be envisioned by substituting NovL with the other three amide synthetases with significant substrate specificity discrepancies.As a major class of natural products, aromatic polyketides display an amazingly creative occupancy of chemical space translating often to their profound bioactivities. Despite the diversity of chemical structures displayed by aromatic polyketides, very few of them contain amide moieties. Several amide-containing aromatic polyketides are shown in Fig. 1. Among them, the amide moiety in oxytetracycline is proposed to originate from a specific malonamyl starter unit (7,8). In the case of lactonamycin, labeling studies suggest that the amino group originates from glycine or a glycine-derived starter unit (9). Fredericamycin (FDM) 2 A (10), pradimicin A (11), CBS40 (12), and lysolipin X (13) are all aromatic polyketides originating from polyketide chains of at least 24 carbons in length. Amide construction in these natural products has not yet received significant attention, and thus the mechanisms associated with these transformations remain obscure.FDM A is a pentadecaketide-derived aromatic compound isolated from Streptomyces griseus ATCC 49344 in 1981 (10, 14). It is notable for its remarkable antitumor potential and its unique carbaspirocyclic structure (15, 16). We previously cloned and sequenced the fdm gene cluster, setting the stage to study the biosynthetic pathway responsible for FDM A production (17). Reexamination of S. griseus wild-type fermentation recently resulted in the isolation of three FDM analogs FDM B, FDM C, and FDM E (18,19) (Fig. 2). FDM B and FDM E both contain the lactam ring F, and FDM C has a carboxylic acid group comparably situated. These facts support the proposal that the amide nitrogen of FDM A is incorporated following installation of the polycyclic ring system. The * This work was su...

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“…Activation of pathwayspecific positive regulatory gene is commonly used for augmentation of secondary metabolites in Streptomyces. [8][9][10] FQs, originally isolated from the culture broth of Streptomyces sp. KO-3988, 11 have cytocidal activities without antibacterial activity.…”

Section: Introductionmentioning

confidence: 99%

Furaquinocins I and J: novel polyketide isoprenoid hybrid compounds from Streptomyces reveromyceticus SN-593

et al. 2011

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Two novel furaquinocin (FQ) analogues, I (1) and J (2), were isolated from Streptomyces reveromyceticus SN-593 strain NRM2. Their structures were elucidated by MS and NMR analyses. Similar to the previously described FQ D (3), both 1 and 2 possessed a dihydrofuran ring fused to a polyketide naphthoquinone skeleton. The main difference between 1, 2 and 3 was the type of residue attached to C-13; these were a carboxyl, a carboxamide and a methyl residue, respectively. INTRODUCTIONThe majority of bioactive compounds used as drugs and bioprobes originally derive from the secondary metabolites of microorganisms. Genome sequence analyses have shown that the number of bioactive compounds isolated from Streptomyces spp. is much lower than the number of gene clusters harbored by each species. [1][2][3] This indicates that the majority of gene clusters are either not expressed, or are only minimally expressed, under standard laboratory conditions. To discover novel natural products that are normally expressed at a very low level, we have been performing a systematic isolation of secondary metabolites. As a result, we isolated verticilactam 4 from Streptomyces spiroverticillatus JC-8444, a tautomycin producer, 5 and 6-dimethylallylindole-3-carbaldehyde 6 from S. reveromyceticus SN-593, a reveromycin producer. 7 In addition, from S. reveromyceticus SN-593, we detected metabolites showing UV spectra similar to furaquinocin (FQ), but different mass from the reported FQs. Consistent with the finding, we could annotate an FQ biosynthetic gene cluster from the genome draft sequence. However, with the quantity produced by wild type strain, it was not possible to isolate and determine the structure. So, in order to improve the production of novel FQ derivatives, we performed the expression of pathway-specific regulatory genes associated with the FQ biosynthetic gene cluster. Activation of pathwayspecific positive regulatory gene is commonly used for augmentation of secondary metabolites in Streptomyces. [8][9][10] FQs, originally isolated from the culture broth of Streptomyces sp. 11 have cytocidal activities without antibacterial activity. So far, eight analogues of these polyketide isoprenoid hybrid compounds (Figure 1) [11][12][13] and their gene cluster 14 have been identified.

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Identification and Utility of FdmR1 as a Streptomyces Antibiotic Regulatory Protein Activator for Fredericamycin Production in Streptomyces griseus ATCC 49344 and Heterologous Hosts

Cited by 83 publications

References 46 publications

Cloning of the biosynthetic gene cluster for naphthoxanthene antibiotic FD-594 from Streptomyces sp. TA-0256

Cloning of the biosynthetic gene cluster for naphthoxanthene antibiotic FD-594 from Streptomyces sp. TA-0256

Characterization of FdmV as an Amide Synthetase for Fredericamycin A Biosynthesis in Streptomyces griseus ATCC 43944

Furaquinocins I and J: novel polyketide isoprenoid hybrid compounds from Streptomyces reveromyceticus SN-593

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