In this study, the authors cloned a glycosyltransferase gene PpUGT2 from Paris polyphylla var. yunnanensis with the ORF length of 1 773 bp and encoding 590 amino acids. The phylogenetic tree revealed that PpUGT2 belonged to the UGT80A subfamily and was named as UGT80A49 by the UDP-glycosyltransferase(UGT) Nomenclature Committee. The expression vector pET28a-PpUGT2 was constructed, and enzyme catalytic reaction in vitro was conducted via inducing protein expression and extraction. With UDP-glucose as sugar donor and diosgenin and pennogenin as substrates, the protein was found with the ability to catalyze the C-3 hydroxyl β-glycosylation of diosgenin and pennogenin. To further explore its catalytic characteristic, 15 substrates including steroids and triterpenes were selected and PpUGT2 showed its activity towards the C-17 position of sterol testosterone with UDP-glucose as sugar donor. Homology modelling and molecule docking of PpUGT2 with substrates predicted the key residues interacting with ligands. The re-levant residues of PpUGT2-ligand binding model were scanned to calculate the corresponding mutants, and the optimized mutants were obtained according to the changes in binding affinity of the ligand with protein and the surrounding residues within 5.0 Å of ligands, which had reference value for design of the mutants. This study laid a foundation for further exploring the biosynthetic pathway of polyphyllin as well as the structure of sterol glycosyltransferases.
Ligands ; Glycosyltransferases/genetics* ; Sterols ; Phylogeny ; Ascomycota ; Liliaceae/chemistry* ; Melanthiaceae ; Diosgenin ; Sugars ; Glucose ; Uridine Diphosphate

The endophytic fungus Nigrospora sphaerica S5 derived from the semi-mangrove plant Myoporum bontioides was fermented. Its metabolites were purified by column chromatography. Nine compounds were obtained and identified as terezine P(1), 3-(1-hydroxyethyl)-4-methyl dihydrofuran-2(3H)-one(2), methylhydroheptelidate(3), hydroheptelidic acid(4), 5, 7-dimethoxy-4, 6-dimethylphthalide(5),(3R,4S)-(-)-4-hydroxymellein(6), pestalopyrone(7), indole-3-formaldehyde(8) and p-hydroxybenzaldehyde(9) by spectroscopic techniques. Terezine P(1) was a new alkaloid belonging to the terezine class with a pyrazine ring. Compounds 2-7 were lactones, of which 3 and 4 belonged to sesquiterpenes. Compounds 8 and 9 were indole alkaloids and phenols, respectively. Compounds 3-6 were purified from Nigrospora sp. for the first time. These compounds showed different degrees of antibacterial activity against Staphylococcus aureus, Escherichia coli of O6 serotype and E. coli of O78 serotype.
Alkaloids ; Anti-Bacterial Agents/pharmacology* ; Ascomycota/chemistry* ; Escherichia coli ; Formaldehyde ; Indoles/pharmacology* ; Lactones ; Molecular Structure ; Myoporum/microbiology* ; Phenols ; Pyrazines ; Sesquiterpenes

Four new diphenyl ethers, named epicoccethers K-N (1-4), were purified from the fermentation medium of a fungus Epicoccum sorghinum derived from Myoporum bontioides, and identified through HR-ESI-MS and NMR spectral analysis. Except that compound 1 showed moderate antifungal activity against Penicillium italicum and Fusarium graminearum, the other three compounds showed stronger activity against them than triadimefon. All of them showed moderate or weak antibacterial activity towards Staphylococcus aureus and Escherichia coli with O6 and O78 serotypes except that 3 was inactive to E. coli O6.
Anti-Bacterial Agents/pharmacology* ; Antifungal Agents/chemistry* ; Ascomycota ; Escherichia coli ; Microbial Sensitivity Tests ; Molecular Structure ; Phenyl Ethers/chemistry*

Using the White as basic medium, the effects of the exogenous IBA and endophytic fungal elicitor on the growth of in vitro roots cultures of Dysosma versipellis and production of podophyllotoxin were investigated in this study. The results showed that the IBA and the endophytic fungus Zasmidium syzygii elicitor could increase the content of podophyllotoxin of in vitro roots of D. versipellis after 3 weeks. The White medium added with 3 mg·L~(-1) IBA induced the highest increase of podophyllotoxin(1 830.86 μg·g~(-1)), which was 2.07 folds greater than the control, and followed by 1.5 mg·L~(-1) IBA, fungal elicitor, 1 mg·L~(-1) IBA, 0.5 mg·L~(-1) IBA and 4.5 mg·L~(-1) IBA, which was 1.82, 1.71, 1.63, 1.43 and 1.1 folds greater than the control, respectively. The results also showed that the growth of roots was certain positively correlated with the change of IBA concentration. Therefore, 3 mg·L~(-1) IBA was the most suitable for the production of podophyllotoxin in the in vitro roots of D. versipellis, and the stimulating effect of Z. syzygii fungal elicitor was between 1.5 mg·L~(-1) and 1 mg·L~(-1) IBA, which was a potential natural elicitor to induce the accumulation of podophyllotoxin in future production.
Ascomycota ; Berberidaceae ; chemistry ; Endophytes ; Plant Roots ; drug effects ; growth & development ; Podophyllotoxin ; biosynthesis ; Tissue Culture Techniques

To search for the active diuretic fractions of Clematidis Armandii Caulis( CAC) and determine its main active chemical components by using liquid chromatography-mass spectrometry( LC-MS) and diuretic activity evaluation. CAC 75% ethanol extracts and extracts from different polar solvents were orally administered to saline-loaded rats at different doses. 6 h urinary volume,p H and contents of electrolyte Na+,K+and Cl-were measured. The chemical components of the active fractions were separated and identified by ultra performance liquid chromatography-electrospray ionization-quadrupole time of flight-mass spectrometry( UPLC-ESI-Q-TOF-MS/MS) method. As compared with the control group,the urine volume was increased by 44%( P< 0. 01) and 34%( P < 0. 05) in CAC75% ethanol extract 57. 74 and 28. 8 mg·kg-1 groups respectively; the Na+excretion was increased by 52%( P< 0. 01) and 45%( P<0. 05),respectively; while the Cl-excretion was increased by 101%( P<0. 01) and 85%( P<0. 05),respectively. The urine volume,Na+excretion and Cl-excretion were increased by 50%( P< 0. 01),58%( P< 0. 05),and 65%( P< 0. 05) respectively in petroleum ether extract 70. 98 mg·kg-1 group as compared with the control group. While for the n-butanol extract 194. 18 mg·kg-1 group,the urine volume,Na+and Cl-excretion were increased by 42%( P<0. 01),41%( P<0. 05) and 97%( P<0. 01),respectively. The diuretic activity of other fractions was not obvious. There was no statistical difference in K+excretion in all groups. The results of LC-MS analysis showed that six compounds,including two sterols,one chromogen and three fatty acids,were identified from petroleum ether extract.Fourteen compounds,including six triterpenoid saponins,six lignin glycosides,one sterol glycoside and one phenolic glycoside,were identified from the n-butanol extract. All the results suggested that the ethanol extract of CAC had remarkable diuretic activity and its main effective components included sterol,triterpenoid saponin and lignin glycosides.
Animals ; Ascomycota ; chemistry ; Diuretics ; pharmacology ; Materia Medica ; pharmacology ; Rats ; Solvents ; Spectrometry, Mass, Electrospray Ionization ; Tandem Mass Spectrometry

A total of 27 endophytic fungal strains were isolated from Huperzia serrata,which were richly distributed in the stems and leaves while less distributed in roots. The 27 strains were identified by Internal Transcribed Spacer( ITS) r DNA molecular method and one of the strains belongs to Basidiomycota phylum,and other 26 stains belong to 26 species,9 general,6 families,5 orders,3 classes of Ascomycota Phylum. The dominant strains were Colletotrichum genus,belonging to Glomerellaceae family,Glomerellales order,Sordariomycetes class,Ascomycota Phylum,with the percentage of 48. 15%. The inhibitory activities of the crude extracts of 27 endophytic fungal strains against acetylcholinesterase( ACh E) and nitric oxide( NO) production were evaluated by Ellman's method and Griess method,respectively. Crude extracts of four fungi exhibited inhibitory activities against ACh E with an IC50 value of 42. 5-62. 4 mg·L~(-1),and some fungi's crude extracts were found to inhibit nitric oxide( NO) production in lipopolysaccharide( LPS)-activated RAW264. 7 macrophage cells with an IC50 value of 2. 2-51. 3 mg·L~(-1),which indicated that these fungi had potential anti-inflammatory activities.The chemical composition of the Et OAc extract of endophytic fungus HS21 was also analyzed by LCMS-IT-TOF. Seventeen compounds including six polyketides,four diphenyl ether derivatives and seven meroterpenoids were putatively identified.
Acetylcholinesterase ; Animals ; Anti-Inflammatory Agents ; isolation & purification ; pharmacology ; Ascomycota ; chemistry ; classification ; isolation & purification ; Cholinesterase Inhibitors ; isolation & purification ; metabolism ; Endophytes ; classification ; isolation & purification ; Huperzia ; microbiology ; Mice ; RAW 264.7 Cells

In order to find new source of antifungal agents, eleven cultivable endophytic fungi were isolated from the roots,stems and leaves of Chelidonium majus by traditional method. Seven of them were identified as Colletotrichum(L1, L2, L3, S1, S3, S4, S5), and three of them were identified as Fusarium(R1,R2,R3) by morphological features and molecular biological technology. The antifungal activity test showed that all the tested fungi displayed some inhibitory activity against five common plant pathogens(C. gloeosporioides, Curvularia lunata, Pyricularia oryza, Alternaria alternate and A. brassicae), and their inhibition rate of some test items were over 60%. Among them, R1, S2, S3 and S4 were more potent than others. This study enriches the understanding of endophytes from Ch. majus and provides a basis for the study of new microbial fungicides.
Alternaria ; pathogenicity ; Antibiosis ; Ascomycota ; pathogenicity ; Chelidonium ; microbiology ; Colletotrichum ; chemistry ; isolation & purification ; Endophytes ; chemistry ; isolation & purification ; Fusarium ; chemistry ; isolation & purification

An extracellular lipase from Aureobasidium pullulans was obtained and purified with a specific activity of 17.7 U/mg of protein using ultrafiltration and a DEAE-Sepharose Fast Flow column. Characterization of the lipase indicated that it is a novel finding from the species A. pullulans. The molecular weight of the lipase was 39.5 kDa, determined by sodium dodecyl sulfonate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme exhibited its optimum activity at 40 °C and pH of 7. It also showed a remarkable stability in some organic solutions (30%, v/v) including n-propanol, isopropanol, dimethyl sulfoxide (DMSO), and hexane. The catalytic activity of the lipase was enhanced by Ca²⁺ and was slightly inhibited by Mn²⁺ and Zn²⁺ at a concentration of 10 mmol/L. The lipase was activated by the anionic surfactant SDS and the non-ionic surfactants Tween 20, Tween 80, and Triton X-100, but it was drastically inhibited by the cationic surfactant cetyl trimethyl ammonium bromide (CTAB). Furthermore, the lipase was able to hydrolyze a wide variety of edible oils, such as peanut oil, corn oil, sunflower seed oil, sesame oil, and olive oil. Our study indicated that the lipase we obtained is a potential biocatalyst for industrial use.
Ascomycota/enzymology* ; Calcium ; Catalysis ; Corn Oil/metabolism* ; Detergents/chemistry* ; Enzyme Stability ; Fungal Proteins/chemistry* ; Glucans/chemistry* ; Hexanes/chemistry* ; Hydrogen-Ion Concentration ; Hydrolysis ; Industrial Microbiology ; Lipase/chemistry* ; Manganese/chemistry* ; Olive Oil/metabolism* ; Peanut Oil/metabolism* ; Sesame Oil/metabolism* ; Substrate Specificity ; Sunflower Oil/metabolism* ; Surface-Active Agents ; Temperature ; Zinc/chemistry*

Phytochemical investigation of the culture of Epicoccum nigrum,an endolichenic fungus inhabiting Leptogium masiaticum,led to the isolation of 11 compounds. Based on NMR spectroscopy and HRESIMS data,their structures were determined as one alkaloid fusaricide( 1),and seven benzofuran derivatives including epicoccone( 2),4,6-dihydroxy-5-methoxy-7-methyl-1,3-dihydro isobenzofuran( 3),5-methyl-epicoccone B( 4),3,6,7-trihydroxy-5-methoxy-4-methylisobenzo furan-1( 3 H)-one( 5),3-methoxyepicoccone B( 6),2,3,4-trihydroxy-6-( hydroxymethyl)-5-methylbenzyl-alcohol( 7),and isoochracinic acid( 8),together with three epicoccolide analogs epicocconigrones A( 9),epicoccolide B( 10),and epicocconigrones B( 11). Compounds 1,9 and 10 showed potent microorganism inhibitory effects. These results indicated the potential perspective of this endophytic fungus as an eco-friendly biocide.
Ascomycota/chemistry* ; Endophytes/chemistry* ; Magnetic Resonance Spectroscopy ; Microbial Sensitivity Tests ; Secondary Metabolism

A new chromene (1) and six known compounds identified as 6-hydroxymellein (2), 6-hydroxy-5-methylmellein (3) nectriapyrone (4), chermesinone A(5), chermesinone B(6), and pomopxanthone A(7), were isolated in our investigation of the cytotoxic constituents from the fermented rice substrate of endophytic fungus Phomopsis sp. HCCB03519. The structures of these com pounds were elucidated through spectroscopic data analysis. All compounds exhibited inhibitory activities against cancer cell lines. Compound 7 showed stronger inhibition against cancer cells than the positive control 5-Fu.
Antineoplastic Agents ; chemistry ; pharmacology ; Ascomycota ; chemistry ; Benzopyrans ; chemistry ; pharmacology ; Cell Line, Tumor ; Fluorouracil ; chemistry ; pharmacology ; Humans ; Isocoumarins ; chemistry ; pharmacology ; Molecular Structure