Discovery of a Novel Analogue of FR901533 and the Corresponding Biosynthetic Gene Cluster From Streptosporangium Roseum No. 79089

FR901533 (1, also known as WS79089B), WS79089A (2), and WS79089C (3) are polycyclic aromatic natural products with promising inhibitory activity to endothelin-converting enzymes. In this work, we isolated five tridecaketide products from Streptosporangium roseum No. 79089, including 1-3, benaphthamycin (4) and a novel FR901533 analogue (5). The structure of 5 was characterized based on spectroscopic data. Compared to the major product 2, the new compound 5 has an additional hydroxyl group at C-12 and an extra methyl group at the 13-OH. The configuration of C-19 of these compounds was determined to be R using Mosher\u27s method. A putative biosynthetic gene cluster for compounds 1-5 was discovered by analyzing the genome of S. roseum No. 79089. This 38.6-kb gene cluster contains 38 open reading frames, including a minimal polyketide synthase (wsaA-C), an aromatase (wsaD), three cyclases (wsaE, F and W) and a series of tailoring enzymes such as monooxygenases (wsaO1-O7) and methyltransferases (wsaM1 and M2). Disruption of the ketosynthase gene (wsaA) in this gene cluster abolished the production of 1-5, confirming that this gene cluster is indeed responsible for the biosynthesis of 1-5. A type II polyketide biosynthetic pathway was proposed for this group of natural endothelin-converting enzyme inhibitors

An Efficient Process for Co-Production of γ-Aminobutyric Acid and Probiotic \u3ci\u3eBacillus subtilis\u3c/i\u3e Cells

This study was to establish an integrated process for the co-production of γ-aminobutyric acid (GABA) and live probiotics. Six probiotic bacteria were screened and Bacillus subtilis ATCC 6051 showed the highest GABA-producing capacity. The optimal temperature and initial pH value for GABA production in B. subtilis were found to be 30 °C and 8.0, respectively. A variety of carbon and nitrogen sources were tested, and potato starch and peptone were the preferred carbon and nitrogen sources for GABA production, respectively. The concentrations of carbon source, nitrogen source and substrate (sodium L-glutamate) were then optimized using the response surface methodology. The GABA titer and concentration of viable cells of B. subtilis reached 19.74 g/L and 6.0 × 108 cfu/mL at 120 h. The GABA titer represents the highest production of GABA in B. subtilis. This work thus demonstrates a highly efficient co-production process for GABA and probiotic B. subtilis cells

Chemical Vapor Deposition at High Pressure in a Microgravity Environment

In this paper we present an evaluation of critical requirements of organometallic chemical vapor deposition (OMCVD) at elevated pressure for a channel flow reactor in a microgravity environment. The objective of using high pressure is to maintain single-phase surface composition for materials that have high thermal decomposition pressure at their optimum growth temperature. Access to microgravity is needed to maintain conditions of laminar flow, which is essential for process analysis. Based on ground based observations we present an optimized reactor design for OMCVD at high pressure and reduced gravity. Also, we discuss non-intrusive real-time optical monitoring of flow dynamics coupled to homogeneous gas phase reactions, transport and surface processes. While suborbital flights may suffice for studies of initial stages of heteroepitaxy experiments in space are essential for a complete evaluation of steady-state growth

Lysine-specific demethylase 5C promotes hepatocellular carcinoma cell invasion through inhibition BMP7 expression

The primers used for the amplification of the indicated genes.(DOCX 17 kb

Engineered Production of Fungal Anticancer Cyclooligomer Depsipeptides in Saccharo-Myces Cerevisiae

Two fungal cyclooligomer depsipeptide synthetases (CODSs), BbBEAS (352 kDa) and BbBSLS (348 kDa) from Beauveria bassiana ATCC 7159, were reconstituted in Saccharomyces cerevisiae BJ5464-NpgA, leading to the production of the corresponding anticancer natural products, beauvericins and bassianolide, respectively. The titers of beauvericins (33.82±1.41 mg/l) and bassianolide (21.74±0.08 mg/l) in the engineered S. cerevisiae BJ5464-NpgA strains were comparable to those in the native producer B. bassiana. Feeding D-hydroxyisovaleric acid (D-Hiv) and the corresponding L-amino acid precursors improved the production of beauvericins and bassianolide. However, the high price of D-Hiv limits its application in large-scale production of these cyclooligomer depsipeptides. Alternatively, we engineered another enzyme, ketoisovalerate reductase (KIVR) from B. bassiana, into S. cerevisiae BJ5464-NpgA for enhanced in situ synthesis of this expensive substrate. Co-expression of BbBEAS and KIVR in the yeast led to significant improvement of the production of beauvericins. The total titer of beauvericin and its congeners (beauvericins A, B and C) was increased to 61.73±2.96 mg/l and reached 2.6-fold of that in the native producer B. bassiana ATCC 7159. Supplement of L-Val at 10 mM improved the supply of ketoisovalerate, the substrate of KIVR, which consequently further increased the total titer of beauvericins to 105.76±2.12 mg/l. Using this yeast system, we functionally characterized an unknown CODS from Fusarium venenatum NRRL 26139 as a beauvericin synthetase, which was named as FvBEAS. Our work thus provides a useful approach for functional reconstitution and engineering of fungal CODSs for efficient production of this family of anticancer molecules

A New Anthracene Derivative from Marine Streptomyces sp. W007 Exhibiting Highly and Selectively Cytotoxic Activities

A new anthracene derivative, 3-hydroxy-1-keto-3-methyl-8-methoxy-1,2,3, 4-tetrahydro-benz[α]anthracene, was isolated from the marine strain Streptomyces sp. W007, and its structure was established by spectroscopic analysis including mass spectra, 1D- and 2D-NMR (1H–1H COSY, HMBC, HSQC and NOESY) experiments. 3-hydroxy-1-keto-3-methyl-8-methoxy-1,2,3,4-tetrahydro-benz[α]anthracene showed cytotoxicity against human lung adenocarcinoma cell line A549

Identification and Heterologous Reconstitution of a 5-Alk(en)ylresorcinol Synthase from Endophytic Fungus \u3ci\u3eShiraia\u3c/i\u3e sp. Slf14

A new type III polyketide synthase gene (Ssars) was discovered from the genome of Shiraia sp. Slf14, an endophytic fungal strain from Huperzia serrata. The intron-free gene was cloned from the cDNA and ligated to two expression vectors pET28a and YEpADH2p-URA3 for expression in Escherichia coli BL21(DE3) and Saccharomyces cerevisiae BJ5464, respectively. SsARS was efficiently expressed in E. coli BL21(DE3), leading to the synthesis of a series of polyketide products. Six major products were isolated from the engineered E. coli and characterized as 1,3-dihydroxyphenyl-5-undecane, 1,3-dihydroxyphenyl-5-cis-6\u27-tridecene,1,3-dihydroxyphenyl-5-tridecane, 1,3-dihydroxyphenyl-5-cis-8\u27-pentadecene, 1,3-dihydroxyphenyl-5-pentadecane and 1,3-dihydroxyphenyl-5-cis-10\u27-heptadecene, respectively, based on the spectral data and biosynthetic origin. Expression of SsARS in the yeast also led to the synthesis of the same polyketide products, indicating that this enzyme can be reconstituted in both heterologous hosts. Supplementation of soybean oil into the culture of E. coli BL21(DE3)/SsARS increased the production titers of 1-6 and led to the synthesis of an additional product, which was identified as 5-(8\u27Z,11\u27Z-heptadecadienyl)resorcinol. This work thus allowed the identification of SsARS as a 5-alk(en)ylresorcinol synthase with flexible substrate specificity toward endogenous and exogenous fatty acids. Desired resorcinol derivatives may be synthesized by supplying corresponding fatty acids into the culture medium

Three-Dimensional Stochastic Distribution Characteristics of Void Fraction in Longwall Mining-Disturbed Overburden of Inclined Coal Seam

AbstractFractures in the overburden induced by mining disturbances provide a channel for fluid flow between the surface and the underground. Mining-induced strata movement and fracture distribution are influenced by the gravity and dip angles of rock seams. In this paper, a new three-dimensional theoretical distribution model for void fraction in each partition of overlying rock strata disturbed by inclined coal seam mining was constructed. Based on the theoretical determination model, the three-dimensional random distribution characteristics for void fraction were obtained by combining the random distribution law of void fraction obtained by similar physical simulation experiments and image processing techniques. Theoretical deterministic models, stochastic theoretical models, and similar physical simulations all show that void fraction distribution in the tendency direction of the coal seam shows a bimodal asymmetric distribution with high and low peaks and a symmetric distribution in the strike direction. The void fraction of the overburden in the central part of the mining area is smaller than that of the surrounding area. The results of the theoretically determined model and stochastic model of the void fraction for the strata with different mining lengths and different coal seam inclinations were compared with the results of similar simulation experiments, respectively. The results are in agreement, further verifying the practicality of the model