RESUMEN
Ganoderma is a well-known genus of medicinal mushrooms. The biological activity of the fruiting bodies of G. mbrekobenum (previously identified as Ganoderma sp. EGDA, (AC: LN774971) is scarcely studied. The microorganisms including bacteria and fungi were chosen for screening of the antimicrobial activity produced by G. mbrekobenum strain EGDA. The bioactive compounds were extracted from aqueous, petroleum ether, chloroform, ethyl acetate, and methanol extracts. The higher antibacterial activity produced by methanol extract was against Bacillus subtilis and B. cereus (14.13 ± 0.12 mm, 13.03 ± 0.12 mm, respectively). Water fraction showed antibacterial effect against most of the test bacterial strains. The highest antifungal activity produced by methanol extract was against Fusarium oxysporum I and F. oxysporum f. sp. lycopersici (16.37 ± 0.03 mm 15.67 ± 0.19 mm, respectively). Gas chromatography/mass spectrometry analysis of the separated fractions revealed the identification of 46 compounds.
Asunto(s)
Ganoderma , Metanol , Egipto , Antifúngicos/farmacología , Antifúngicos/química , Antibacterianos/farmacología , Antibacterianos/química , Extractos Vegetales/química , Bacillus subtilisRESUMEN
Ergosterol peroxide and ganoderic acid AMI were isolated for the first time from the mycelium of the Egyptian Ganoderma resinaceum mushroom. The structure of these two metabolites was established by detailed analysis of 1D and 2D NMR. The isolated compounds were tested for their antitumor in vitro activities in MCF-7 and MDA-MB-231 breast cancer cell lines. Ergosterol peroxide showed preferred inhibition of MCF-7 (ER +ve) cell lines relative to MDA-MB-231 (ER -ve) cell lines with an IC50 of 1.18 µM and 12.82 µM respectively. Our data suggest that ergosterol peroxide targets estrogen receptors.
Asunto(s)
Antineoplásicos/farmacología , Ergosterol/análogos & derivados , Ganoderma/química , Antineoplásicos/química , Antineoplásicos/aislamiento & purificación , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Egipto , Ergosterol/química , Ergosterol/aislamiento & purificación , Ergosterol/farmacología , Humanos , Concentración 50 Inhibidora , Células MCF-7 , Estructura Molecular , Micelio/química , Receptores de Estrógenos/metabolismo , Triterpenos/química , Triterpenos/aislamiento & purificación , Triterpenos/farmacologíaRESUMEN
AmyLa α-amylase gene from Laceyella sp. DS3 was heterologously expressed in E. coli BL21. E. coli BL21 maximally expressed AmyLa after 4â¯h of adding 0.02â¯mM IPTG at 37⯰C. The recombinant AmyLa α-amylase was purified 2.19-fold through gel filtration and ion exchange chromatography. We immobilized the purified recombinant AmyLa α-amylase on four carriers; chitosan had the best efficiency. The recombinant free and the immobilized AmyLa α-amylase showed optimum activity in the pH ranges of 6.0-7.0 and 4.0-7.0, respectively and possessed an optimum temperature of 55⯰C. The free enzyme had activation energy, Km, and Vmax of 291.5â¯kJ, 1.5â¯mg/ml, and 6.06â¯mg/min, respectively. The immobilized enzyme had activation energy, Km, and Vmax of 309.74â¯kJ, 6.67â¯mg/ml, and 50â¯mg/min, respectively. The immobilized enzyme was calcium-independent and insensitive (relative to the free enzyme) to metals. It could also be reused for seven cycles.
Asunto(s)
Bacillales/enzimología , Enzimas Inmovilizadas/genética , Enzimas Inmovilizadas/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , alfa-Amilasas/genética , alfa-Amilasas/metabolismo , Enzimas Inmovilizadas/química , Enzimas Inmovilizadas/aislamiento & purificación , Escherichia coli/genética , Expresión Génica , Cinética , Proteínas Recombinantes/química , Proteínas Recombinantes/aislamiento & purificación , alfa-Amilasas/química , alfa-Amilasas/aislamiento & purificaciónRESUMEN
α-Amylase from Thermoactinomyces vulgaris was highly purified 48.9-fold by ammonium sulfate precipitation, gel filtration on Sephadex G-50 column, and ion exchange chromatography column of DEAE-cellulose. The molecular weight of the enzyme was estimated to be 135 and 145 kDa by SDS-PAGE. Its high molecular weight is due to high glycosylation. The purified amylase exhibited maximal activity at pH 6.0 to 7.0 and was stable in the range of pH 4.0 to 9.0. The optimum temperature for its activity was 50 °C. The enzyme half-life time was 120 min at 50 °C, suggesting intermediate temperature stable α-amylase. The enzyme was sensitive to different metal ions, including NaCl, CoCl(2), and CaCl(2), and to different concentrations of EDTA. The enzyme activity was inhibited in the presence of 1 mM CaCl(2), suggesting that it is a calcium-independent α-amylase. The TLC showed that the amylase hydrolyzed starch to produce large maltooligosaccharides as the main products. A 1.1-kb DNA fragment of the putative α-amylase gene (amy TVE) from T. vulgaris was amplified by using two specific newly designed primers. Sequencing analysis showed 56.2 % similarity to other Thermoactinomyces α-amylases with two conserved active sites confirming its function.