Streptomyces are a genus of ubiquitous soil bacteria from which the majority of clinically utilized antibiotics derive
. The production of these antibacterial molecules reflects the relentless ...competition Streptomyces engage in with other bacteria, including other Streptomyces species
. Here we show that in addition to small-molecule antibiotics, Streptomyces produce and secrete antibacterial protein complexes that feature a large, degenerate repeat-containing polymorphic toxin protein. A cryo-electron microscopy structure of these particles reveals an extended stalk topped by a ringed crown comprising the toxin repeats scaffolding five lectin-tipped spokes, which led us to name them umbrella particles. Streptomyces coelicolor encodes three umbrella particles with distinct toxin and lectin composition. Notably, supernatant containing these toxins specifically and potently inhibits the growth of select Streptomyces species from among a diverse collection of bacteria screened. For one target, Streptomyces griseus, inhibition relies on a single toxin and that intoxication manifests as rapid cessation of vegetative hyphal growth. Our data show that Streptomyces umbrella particles mediate competition among vegetative mycelia of related species, a function distinct from small-molecule antibiotics, which are produced at the onset of reproductive growth and act broadly
. Sequence analyses suggest that this role of umbrella particles extends beyond Streptomyces, as we identified umbrella loci in nearly 1,000 species across Actinobacteria.
The main aim of the current study is to explore the bioactive potential of Streptomyces sp. VITJS8 isolated from the marine saltern. The cultural, biochemical, and morphological studies were ...performed to acquire the characteristic features of the potent isolate VITJS8. The 16Sr DNA sequencing was performed to investigate the phylogenetic relationship between the Streptomyces genera. The structure of the compound was elucidated by gas chromatography-mass spectrometry (GC-MS), infra-red (IR), and ultra-violet (UV) spectroscopic data analysis. The GC-MS showed the retention time at 22.39 with a single peak indicating the purity of the active compound, and the molecular formula was established as C sub(14)H sub(9)ONCl sub(2 ) based on the peak at m/z 277 M super(+). Furthermore, separated by high-performance liquid chromatography (HPLC), their retention time (t sub(r)) 2.761 was observed with the absorption maxima at 310 nm. The active compound showed effective inhibitory potential against four clinical pathogens at 500 mu g/mL. The antioxidant activity was found effective at the IC sub(50) value of 500 mu g/mL with 90 % inhibition. The 3-(4,5-dimethylthiazol-2-yl)-2,5-ditetrazolium bromide (MTT) assay revealed the cytotoxicity against HepG2 cells at IC sub(50) of 250 mu g/mL. The progression of apoptosis was evidenced by morphological changes by nuclear staining. The DNA fragmentation pattern was observed at 250 mu g/mL concentration. Based on flow cytometric analysis, it was evident that the compound was effective in inhibiting the sub-G0/G1 phase of cell cycle. The in vitro findings were also supported by the binding mode molecular docking studies. The active compound revealed minimum binding energy of -7.84 and showed good affinity towards the active region of topoisomerase-2 alpha that could be considered as a suitable inhibitor. Lastly, we performed 30 ns molecular dynamic simulation analysis using GROMACS to aid in better designing of anticancer drugs. Simulation result of root mean square deviation (RMSD) analysis showed that protein-ligand complex reaches equilibration state around 10 ns that illustrates the docked complex is stable. We propose the possible mechanism of sesquiterpenes to play a significant role in antitumor cascade. Hence, our studies open up a new facet for a potent drug as an anticancer agent.
Laccases (EC 1.10.3.2) are multicopper oxidases that can oxidize a range of substrates, including phenols, aromatic amines, and nonphenolic substrates. To investigate the involvement of the small ...Streptomyces laccases in lignin degradation, we generated acid-precipitable polymeric lignin obtained in the presence of wild-type Streptomyces coelicolor A3(2) (SCWT) and its laccase-less mutant (SCΔLAC) in the presence of Miscanthus x giganteus lignocellulose. The results showed that strain SCΔLAC was inefficient in degrading lignin compared to strain SCWT, thereby supporting the importance of laccase for lignin degradation by S. coelicolor A3(2). We also studied the lignin degradation activity of laccases from S. coelicolor A3(2), Streptomyces lividans TK24, Streptomyces viridosporus T7A, and Amycolatopsis sp. 75iv2 using both lignin model compounds and ethanosolv lignin. All four laccases degraded a phenolic model compound (LM-OH) but were able to oxidize a nonphenolic model compound only in the presence of redox mediators. Their activities are highest at pH 8.0 with a low krel/Kapp for LM-OH, suggesting that the enzymes’ natural substrates must be different in shape or chemical nature. Crystal structures of the laccases from S. viridosporus T7A (SVLAC) and Amycolatopsis sp. 75iv2 were determined both with and without bound substrate. This is the first report of a crystal structure for any laccase bound to a nonphenolic β-O-4 lignin model compound. An additional zinc metal binding site in SVLAC was also identified. The ability to oxidize and/or rearrange ethanosolv lignin provides further evidence of the utility of laccase activity for lignin degradation and/or modification.
Background: Bioactive metabolites that inhibit cellular migration are considered to be useful for the suppression of cancer metastasis. Then, we have looked for cancer cell migration inhibitors from ...microorganisms. In one hand, ovarian carcinoma is highly invasive and often metastasizes into liver, lung, and peritoneal cavity.Materials and methods: Cellular migration was assessed by wound healing assay with breast carcinoma MDA-MB-231 cells. We employed about 1500 microbial broths for random screening. From the culture filtrate with positive activity, active principle was isolated by chromatography. The structure was determined mainly by NMR and mass spectroscopy after the purification.Results: After testing the thousands of microbial culture filtrates, we have isolated novel compounds migracin A and B from Streptomyces. Migracin A and B showed similar inhibitory activity on the migration of MDA-MB-231 cells and fibrosarcoma HT-1080 cells. They also inhibited TNF-alpha and TGF-beta-induced migration of lung adenocarcinoma A549 cells. Migracin A inhibited the Matrigel invasion of clear cell ovarian carcinoma ES-2 cells without any toxicity. Since the structure of migracin is related to that of luminacin that inhibits tube formation, we have studied the effect on lumina formation. As a result, migracin A inhibited VEGF-induced limina formation in HUVEC. The mechanism of inhibition is being studied.Conclusions: We have discovered migracins as novel inhibitors of cellular migration and invasion. Migracin inhibited invasion of ovarian carcinoma cells, and is considered to be a candidate of new metastasis inhibitor.
The roles of many sigma factors are unclear in regulatory mechanism of secondary metabolism in Streptomyces. Here, we report the regulation network of a group 3 sigma factor, WhiG sub(ch), from a ...natamycin industrial strain Streptomyces chattanoogensis L10. WhiG sub(ch) regulates the growth and morphological differentiation of S. chattanoogensis L10. The whiG sub(ch) deletion mutant decreased natamycin production by about 30 % and delayed natamycin production more than 24 h by delaying the growth. Overexpression of the whiG sub(ch) gene increased natamycin production in large scale production medium by about 26 %. WhiG sub(ch) upregulated the transcription of natamycin biosynthetic gene cluster and inhibited the expression of migrastatin and jadomycin analog biosynthetic polyketide synthase genes. WhiG sub(ch) positively regulated natamycin biosynthetic gene cluster by directly binding to the promoters of scnC and scnD, which were involved in natamycin biosynthesis, and these binding sites adjacent to translation start codon were determined. Thus, this paper further elucidates the high natamycin yield mechanisms of industrial strains and demonstrates that a valuable improvement in the yield of the target metabolites can be achieved through manipulating the transcription regulators.
Nucleocidin is one of the very few natural products known to contain fluorine. Mysteriously, the nucleocidin producer Streptomyces calvus ATCC13382 has not been observed to synthesize the compound ...since its discovery in 1956. Here, we report that complementation of S. calvus ATCC13382 with a functional bldA-encoded Leu-tRNA super(UUA) molecule restores the production of nucleocidin. Nucleocidin was detected in culture extracts by super(19)FNMR spectroscopy, HPLC-ESI-MS, and HPLC-continuum source molecular absorption spectroscopy for fluorine-specific detection. The molecule was purified from a large-scale culture and definitively characterized by NMR spectroscopy and high-resolution MS. The nucleocidin biosynthetic gene cluster was identified by the presence of genes encoding the 5'-O-sulfamate moiety and confirmed by gene disruption. Two of the genes within the nucleocidin biosynthetic gene cluster contain TTA codons, thus explaining the dependence on bldA and resolving a 60-year-old mystery. Missing since 1956: The biosynthesis of nucleocidin in Streptomyces calvus has been restored by complementation with a functional bldA-encoded Leu-tRNA super(UUA) molecule. Nucleocidin was detected in culture extracts, purified, and characterized. Its biosynthetic gene cluster was identified and confirmed by gene disruption.
Streptomyces sp. Tue 6176, producer of cytotoxic benzoxazoles AJI9561, nataxazole, and 5-hydroxy-nataxazole, has been found to produce a fourth benzoxazole, UK-1. All derive from ...3-hydroxy-anthranilate synthesized by the nataxazole biosynthesis machinery. However, biosynthesis of AJI9561, nataxazole, and 5-hydroxy-nataxazole requires 6-methylsalicylic acid also provided by nataxazole biosynthesis pathway, while biosynthesis of UK-1 utilizes salicylic acid produced by a salicylate synthase from the coelibactin biosynthesis pathway. This clearly suggests crosstalk between nataxazole and coelibactin pathways. Overproduction of UK-1 was obtained by growing a nataxazole non-producing mutant (lacking 6-methylsalicylate synthase, NatPK) in a zinc-deficient medium. Furthermore, Streptomyces sp. Tue 6176 also produces the siderophore enterobactin in an iron-free medium. Enterobactin production can be induced in an iron-independent manner by inactivating natAN, which encodes an anthranilate synthase involved in nataxazole production. The results indicate a close relationship between nataxazole, enterobactin and coelibactin pathways through the shikimate pathway, the source of their common precursor, chorismate. In- Streptomyces- sp. Tue 6176 there is a close relationship between the biosynthesis pathways of nataxazole and of ionophores enterobactin and coelibactin through the shikimate pathway (source of the common precursor chorismate). In addition, crosstalk between nataxazole and coelibactin pathways is responsible for the biosynthesis of benzoxazole UK-1.
Aims
To evaluate the potential of chitinolytic endophytic Actinomycetes isolated from medicinal plants in order to diminish the collar rot infestation induced by Sclerotium rolfsii in chickpea.
...Methods and Results
Sixty‐eight chitinolytic endophytic Actinomycetes were recovered from various medicinal plants and evaluated for their chitinase activity. Among these isolates, 12 were screened for their plant growth promoting abilities and antagonistic potential against Sc. rolfsii. Further, these isolates were validated in vivo for their ability to protect chickpea against Sc. rolfsii infestation under greenhouse conditions. The isolates significantly (P < 0·05) increased the biomass (1·2–2·0 fold) and reduced plant mortality (42–75%) of chickpea. On the basis of 16S rDNA profiling, the selected antagonistic strains were identified as Streptomyces diastaticus, Streptomyces fradiae, Streptomyces olivochromogenes, Streptomyces collinus, Streptomyces ossamyceticus and Streptomyces griseus.
Conclusion
This study is the first report of the isolation of endophytic Actinomycetes from various medicinal plants having antagonistic and plant growth promoting abilities. The isolated species showed potential for controlling collar rot disease on chickpea and could be useful in integrated control against diverse soil borne plant pathogens.
Significance and Impact of the Study
Our investigation suggests that endophytic Actinomycetes associated with medicinal plants can be used as bioinoculants for developing safe, efficacious and environment‐friendly biocontrol strategies in the near future.
Streptomyces sp. Tue 6176 produces the cytotoxic benzoxazole nataxazole. Bioinformatic analysis of the genome of this organism predicts the presence of 38 putative secondary-metabolite biosynthesis ...gene clusters, including those involved in the biosynthesis of AJI9561 and its derivative nataxazole, the antibiotic hygromycin B, and ionophores enterobactin and coelibactin. The nataxazole biosynthesis gene cluster was identified and characterized: it lacks the O-methyltransferase gene required to convert AJI9561 into nataxazole. This O-methyltransferase activity might act as a resistance mechanism, as AJI9561 shows antibiotic activity whereas nataxazole is inactive. Moreover, heterologous expression of the nataxazole biosynthesis gene cluster in S. lividans JT46 resulted in the production of AJI9561. Nataxazole biosynthesis requires the shikimate pathway to generate 3-hydroxyanthranilate and an iterative type I PKS to generate 6-methylsalicylate. Production of nataxazole was improved up to fourfold by disrupting one regulatory gene in the cluster. An additional benzoxazole, 5-hydroxynataxazole is produced by Streptomyces sp. Tue 6176. 5-Hydroxynataxazole derives from nataxazole by the activity of an as yet unidentified oxygenase; this implies cross-talk between the nataxazole biosynthesis pathway and an unknown pathway. Cross-talk between pathways:- Streptomyces sp. Tue 6176 produces cytotoxic benzoxazole nataxazole, its derivative 5-hydroxynataxazole, and under certain conditions the nataxazole intermediate AJI9561. Biosynthesis uses 3-hydroxyanthranilate from the shikimate pathway and an iterative type I PKS to generate 6-methylsalicylate. Heterologous expression of the gene cluster led to the production of AJI9561.
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