RESUMO
The fungus Fusarium graminearum causes a devastating disease Gibberella stalk rot of maize. Our knowledge of molecular interactions between F. graminearum effectors and maize immunity factors is lacking. Here, we show that a group of cysteine-rich common in fungal extracellular membrane (CFEM) domain proteins of F. graminearum are required for full virulence in maize stalk infection and that they interact with two secreted maize proteins, ZmLRR5 and ZmWAK17ET. ZmWAK17ET is an alternative splicing isoform of a wall-associated kinase ZmWAK17. Both ZmLRR5 and ZmWAK17ET interact with the extracellular domain of ZmWAK17. Transgenic maize overexpressing ZmWAK17 shows increased resistance to F. graminearum, while ZmWAK17 mutants exhibit enhanced susceptibility to F. graminearum. Transient expression of ZmWAK17 in Nicotiana benthamiana triggers hypersensitive cell death, whereas co-expression of CFEMs with ZmWAK17ET or ZmLRR5 suppresses the ZmWAK17-triggered cell death. Our results show that ZmWAK17 mediates stalk rot resistance and that F. graminearum delivers apoplastic CFEMs to compromise ZmWAK17-mediated resistance.
Assuntos
Gibberella , Zea mays , Zea mays/genética , Zea mays/metabolismo , Gibberella/metabolismo , Doenças das Plantas/microbiologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismoRESUMO
The pathogenic fungi Gibberella fujikuroi and Fusarium commune produce jasmonic acid. The application of volatile deuterium-labeled methyl jasmonate increased the amount of nonlabeled JA present in G. fujikuroi and F. commune. These results indicate that the fungi have the ability to react with airborne methyl jasmonate in a manner similar to a plant.
Assuntos
Acetatos/metabolismo , Ciclopentanos/metabolismo , Fusarium/metabolismo , Gibberella/metabolismo , Oxilipinas/metabolismo , Plantas/metabolismo , Poluentes Atmosféricos/metabolismoRESUMO
BACKGROUND: Eugenol is an economically favorable substrate for the microbial biotransformation of aromatic compounds. Coniferyl aldehyde is one kind of aromatic compound that is widely used in condiment and medical industries; it is also an important raw material for producing other valuable products such as vanillin and protocatechuic acid. However, in most eugenol biotransformation processes, only a trace amount of coniferyl aldehyde is detected, thus making these processes economically unattractive. As a result, an investigation of new strains with the capability of producing more coniferyl aldehyde from eugenol is required. RESULTS: We screened a novel strain of Gibberella fujikuroi, labeled as ZH-34, which was capable of transforming eugenol to coniferyl aldehyde. The metabolic pathway was analyzed by high-performance liquid chromatographymass spectrometry and transformation kinetics. The culture medium and biotransformation conditions were optimized. At a 6 h time interval of eugenol fed-batch strategy, 3.76 ± 0.22 g/L coniferyl aldehyde was obtained, with the corresponding yield of 57.3%. CONCLUSIONS: This work improves the yield of coniferyl aldehyde with a biotechnological approach. Moreover, the fed-batch strategy offers possibility for controlling the target product and accumulating different metabolites
Assuntos
Acroleína/análogos & derivados , Eugenol/metabolismo , Biotransformação , Gibberella/metabolismo , Biodegradação Ambiental , Acroleína/metabolismo , Biotecnologia , Cromatografia Líquida de Alta Pressão , Recursos Renováveis , Técnicas de Cultura Celular por LotesRESUMO
Terpenylated phenols possess interesting biological activities. These properties vary mainly according to the type of terpene associated and the degree of oxidation of the molecule. The search for new active molecules for application in different areas of knowledge includes the structural modification of these through ecological methodologies, such as biotransformation. The aims of this study were the biotransformation of geranylated- and acetylated-phloroglucinol by the fungus Gibberella fujikuroi and the evaluation of the antifungal activity of the derivatives. Five major derivatives were identified after biotransformation, highlighting the formation of specific monoacetylated products. In vitro antifungal activity assays against the phytopathogenic fungus Botrytis cinerea indicated that deacetylated derivatives possess higher activity compared to the precursor molecule. In other biotransformation reactions, a relationship between the release of the alkyl chain from the aromatic ring with a decrease of the antifungal activity, was observed. The in vivo tests in infected tomato plants with B. cinerea confirmed the antifungal activity of the derivatives observed in in vitro experiments.
Assuntos
Antifúngicos/metabolismo , Botrytis/efeitos dos fármacos , Gibberella/metabolismo , Floroglucinol/metabolismo , Biotransformação , Solanum lycopersicum/microbiologia , Testes de Sensibilidade Microbiana , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controleRESUMO
Three new (6, 9, and 12) and nine known steroidal saponins were obtained from the fermentation broth of pseudoprotodioscin (PPD) incubated with a fungus Gibberella fujikuroi CGMCC 3.4663. Structures of the metabolites were elucidated by 1-D (1H, 13C), 2-D (HMBC, HSQC, NOESY) NMR, and HR-MS analyses. The biotransformation pathway of pseudoprotodioscin by Gibberella fujikuroi CGMCC 3.4663 was proposed. Compounds 1-11 were tested in vitro for their cytotoxic activities against two human cancer cell lines (HepG2 and Hela). Compounds 1, 6, 9, and 10 exhibited cytotoxic activity against HepG2 cells. Compound 10 exhibited cytotoxicity to Hela cells.
Assuntos
Diosgenina/análogos & derivados , Gibberella/metabolismo , Antibióticos Antineoplásicos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Diosgenina/química , Diosgenina/metabolismo , Diosgenina/farmacologia , Gibberella/química , Células HeLa , Células Hep G2 , Humanos , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Estrutura MolecularRESUMO
Derivatives of spironolactone (1), a diuretic and antihypertensive drug, were synthesized by using fungal cells for the first time. Ten different fungi were screened for their ability to biotransform 1, four of which were able to produce metabolites 2-8. Gibberella fujikuroi produced canrenone (2), 1-dehydrocanrenone (3), Curvularia lunuta provided compound 2, and 7α-thio-spironolactone (4), Fusarium lini yielded compounds 2, 3, 1ß-hydroxycanrenone (5), 1α-hydroxycanrenone (6), 1-dehydro-15α-hydroxycanrenone (7), and 15α-hydroxycanrenone (8), while Aspergillus alliaceus was able to produce all the seven metabolites. Metabolites 5, 6, and 7 were identified as new compounds. Their structures were elucidated by using different spectroscopic techniques. Substrate 1 and its metabolites 2, 3, and 5-8 were also evaluated for α-glucosidase inhibitory activity in vitro. Substrate 1 was found to be strongly active with IC50â¯=â¯335⯱â¯4.3⯵M as compared to the standard drug acarbose IC50â¯=â¯840⯱â¯1.73⯵M, whereas all of resulting metabolites were found to be inactive.
Assuntos
Anti-Hipertensivos/síntese química , Biotransformação , Diuréticos/síntese química , Espironolactona/síntese química , Anti-Hipertensivos/química , Anti-Hipertensivos/uso terapêutico , Ascomicetos/química , Ascomicetos/genética , Ascomicetos/metabolismo , Aspergillus/química , Aspergillus/genética , Aspergillus/metabolismo , Diuréticos/química , Diuréticos/uso terapêutico , Fusarium/química , Fusarium/genética , Fusarium/metabolismo , Gibberella/química , Gibberella/genética , Gibberella/metabolismo , Humanos , Estrutura Molecular , Espironolactona/análogos & derivados , Espironolactona/química , Espironolactona/uso terapêuticoRESUMO
Lawsone (2-hydroxy-1, 4-napthoquinone), also known as hennotannic acid, is an orange red dye used as a popular skin and hair colorant. The dye is produced in the leaves of Lawsonia inermis L, often referred to as the "henna" tree. In this study, we report the production of lawsone by an endophytic fungus, Gibberella moniliformis isolated from the leaf tissues of Lawsonia inermis. The fungus produced the orange-red dye in potato dextrose agar and broth, independent of the host tissue. Presence of lawsone was confirmed spectrometrically using HPLC and ESI-MS/MS analysis. The fragmentation pattern of lawsone was identical to both standard lawsone and that extracted from plant tissue. This is a first report of lawsone being produced by an endophytic fungus, independent of the host tissue. The study opens up interesting questions on the possible biosynthetic pathway through which lawsone is produced by the fungus.
Assuntos
Gibberella/metabolismo , Lawsonia (Planta) , Naftoquinonas/metabolismo , Animais , Citrus sinensis , Fungos , Extratos Vegetais , Espectrometria de Massas em TandemRESUMO
Structural transformation of anticancer drug exemestane (1) with fungi Cunninghamella blakesleeana (ATCC 8688A), Curvularia lunata (ATCC 12017), Aspergillus niger (ATCC 10549), and Gibberella fujikuroi (ATCC 10704) yielded eleven metabolites 2-12, in which 2 and 8 were identified as new. Their structures were characterized as 6-methylene-5α-androstane-3ß,16ß,17ß-triol (2), 17ß-hydroxy-6-methyleneandrosta-4-ene-3-one (3), 6α-spiroxirandrost-4-ene-3,17-dione (4), 6-methyleneandrosta-4-ene-3,17-dione (5), 6ß,17ß-dihydroxyandrost-4-en-3-one (6), 17ß-hydroxy-6α-spiroxirandrost-1,4-diene-3-one (7), 17ß-hydroxy-6α-hydroxymethylandrosta-1,4-dien-3-one (8), 6α-hydroxymethylandrosta-1,4-diene-3,17-dione (9), 17ß-hydroxy-6-methyleneandrosta-1,4-diene-3,16-dione (10), 6α-hydroxy-4-androstene-3,17-dione (11), and 6α-hydroxymethylandrost-4-ene-3,17-dione (12). Substrate 1, and its transformed products were evaluated for their cytotoxicity against breast cancer cell line (MCF-7). Compound 3 was found to be moderately active with an IC50 of 33.43±4.01µM, in comparison to the standard anti-cancer drug, doxorubicin (IC50=0.92±0.1µM).
Assuntos
Androstadienos/metabolismo , Androstadienos/farmacologia , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Neoplasias da Mama/metabolismo , Androstadienos/química , Antineoplásicos/química , Aspergillus niger/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Cunninghamella/metabolismo , Fermentação , Gibberella/metabolismo , Humanos , Células MCF-7 , Estrutura MolecularRESUMO
Microbial transformation of 6-dehydroprogesterone (1) with Aspergillus niger yielded three new metabolites, including 6ß-chloro-7α,11α-dihydroxypregna-4-ene-3,20-dione (2), 7α-chloro-6ß,11α-dihydroxypregna-4-ene-3,20-dione (3), and 6α,7α-epoxy-11α-hydroxypregna-4-ene-3,20-dione (4), and two known metabolites; 6α,7α-epoxypregna-4-ene-3,20-dione (5), and 11α-hydroxypregna-4,6-diene-3,20-dione (6). Compounds 2, and 3 contain chlorohydrin moiety at C-6, and C-7, respectively. The biotransformation of 1 with Gibberella fujikuroi yielded a known compound, 11α,17ß-dihydroxyandrosta-4,6-dien-3-one (7).
Assuntos
Aspergillus niger/metabolismo , Gibberella/metabolismo , Progesterona/química , Progesterona/metabolismo , Biotransformação , Estrutura Molecular , EstereoisomerismoRESUMO
The regio- and stereo-selective hydroxylations of two ingenane diterpenoids, 20-deoxyingenol (1) and 13-oxyingenol dodecanoat (2), by the filamentous fungi Mortierella ramanniana and Gibberella fujikuroi were investigated in the present study. Four undescribed metabolites (3-6) of substrate 1 and two undescribed metabolites (7 and 8) of substrate 2 were isolated. All the metabolites were identified as hydroxylated ingenane derivatives by extensive NMR and HR-ESI-MS data analyses. All the biotransformed compounds and the substrates were evaluated for their cytotoxicities against three human cancer cell lines, including human colon cancer Caco-2, breast cancer MCF-7, and adriamycin (ADM)-resistant MCF-7/ADM cell lines. All ingenane alcohols (1, and 3-6) displayed no significant cytotoxic activities. The substrate 13-oxyingenol dodecanoat (2) showed moderate cytotoxicity with IC50 values being 35.59 ± 5.37 µmol·L-1 (Caco-2), 24.04 ± 4.70 µmol·L-1 (MCF-7), and 22.24 ± 5.19 µmol·L-1 (MCF-7/ADM). However, metabolites 7 and 8 displayed no significant cytotoxicity. These results indicated that the hydroxylation at the C-13 aliphatic acid ester of substrate 2 can significantly reduce the cytotoxic activity.
Assuntos
Diterpenos/química , Diterpenos/metabolismo , Gibberella/metabolismo , Mortierella/metabolismo , Antineoplásicos/química , Antineoplásicos/metabolismo , Biotransformação , Linhagem Celular Tumoral , Humanos , Hidroxilação , Estrutura Molecular , EstereoisomerismoRESUMO
Anti-glycation and α-glucosidase inhibition activities of microbial transformed compounds of dydrogesterone (1); 20R-hydroxy-9ß,10αa-pregna-4,6-diene-3-one (2), 17ß-hydroxy-9ß,10α-androsta-4,6-diene-3-one (3) and 9ß,10α-androsta-4,6-diene-3,17-dione (4) were evaluated. Compounds 1 and 4 showed potent α-glucosidase inhibitory activities, while 2 and 3 were found to be weak inhibitors, whereas anti-glycation activities of 1-4 were not observed.
Assuntos
Didrogesterona/farmacologia , Inibidores de Glicosídeo Hidrolases/farmacologia , Gibberella/metabolismo , Glicosilação/efeitos dos fármacos , Rutina/farmacologiaRESUMO
The aim of this study was to investigate the influence of prebiotic compounds (cellulose and inulin), food ingredients (milk whey, ß-lactoglobulin and calcium caseinate) and several probiotic microorganisms on the bioaccessibility of beauvericin (BEA), enniatins (ENs A, A1, B, B1), deoxynivalenol (DON) and zearalenone (ZEA) present in wheat crispy bread produced with wheat flour previously fermented with F. tricinctum, F. culmorum and G. zeae. The bioaccessibility of mycotoxins was determined by a dynamic simulated gastrointestinal digestion system, imitating the human digestive physiological conditions of the gastrointestinal tract. Mycotoxins were determined in the simulated intestinal fluids by liquid chromatography-tandem mass spectrometry (LC-MS/MS). EN bioaccessibility ranged from 15.1 to 30.6%, whereas the values evidenced for BEA ranged from 12 to 19%. DON showed bioaccessibility data ranging from 0.8 to 5.6% whereas for ZEA the data evidenced ranged from 26 to 44%. The bioaccessibility reduction evidenced using probiotic microorganisms for the mycotoxins studied ranged from 21 to 27.1% for ENs, from 29 to 39.7% for DON, from 41 to 57% for ZEA and from 6.6 to 10.5% for BEA. The addition of prebiotic and bioactive microorganisms decreased the bioaccessibility of mycotoxins, with a concentration-dependent behavior, thus being a potential strategy for reducing human exposure to these minor mycotoxins.
Assuntos
Proteínas Alimentares/uso terapêutico , Ergotismo/prevenção & controle , Trato Gastrointestinal/metabolismo , Modelos Biológicos , Micotoxinas/antagonistas & inibidores , Prebióticos/administração & dosagem , Probióticos/uso terapêutico , Antídotos/uso terapêutico , Bifidobacterium/crescimento & desenvolvimento , Bifidobacterium/metabolismo , Disponibilidade Biológica , Pão/análise , Digestão , Fermentação , Farinha/análise , Farinha/microbiologia , Contaminação de Alimentos , Fusarium/crescimento & desenvolvimento , Fusarium/metabolismo , Conteúdo Gastrointestinal/química , Conteúdo Gastrointestinal/microbiologia , Trato Gastrointestinal/microbiologia , Gibberella/crescimento & desenvolvimento , Gibberella/metabolismo , Humanos , Lactobacillus/crescimento & desenvolvimento , Lactobacillus/metabolismo , Micotoxinas/análise , Micotoxinas/isolamento & purificação , Micotoxinas/toxicidade , Venenos/análise , Venenos/isolamento & purificação , Venenos/toxicidade , ToxicocinéticaRESUMO
The biotransformation of dehydroepiandrosterone (DHEA) by Gibberella intermedia was investigated. The formation of the main products was monitored by high-performance liquid chromatography (HPLC). Products were purified and identified as 3ß,7α-dihydroxy-5-androsten-17-one (7α-OH-DHEA) and 3ß,7α,15α-trihydroxy-5-androsten-17-one (7α,15α-diOH-DHEA) using mass spectroscopy (MS) and nuclear magnetic resonance (NMR) analyses. 7α,15α-DiOH-DHEA is a key intermediate for the synthesis of pharmacologically active steroids. A biotransformation process was optimized to obtain a high concentration of 7α,15α-diOH-DHEA. The results showed the optimal biotransformation process under the following conditions: a culture medium containing 15 g/L glucose, 16 g/L yeast extract, 12 g/L corn steep liquor, and 0.15 g/L ferrous sulfate; an initial pH of 6.5; culture temperature of 30 °C; inoculum size of 4 %; 40 mL volume of the culture medium in 250 mL flasks; the addition of 1 % Tween 80 as co-solvent; and the transformation period of 78 h. After optimization, the molar yield of 7α,15α-diOH-DHEA from 5 g/L substrate DHEA was 77.4 %, which was increased by 115 % than the yield obtained in the original bioconversion process.
Assuntos
Desidroepiandrosterona/metabolismo , Gibberella/metabolismo , Biotransformação/fisiologia , Desidroepiandrosterona/química , Espectroscopia de Ressonância Magnética , Espectrometria de MassasRESUMO
Dihydroxylation of dehydroepiandrosterone (DHEA) is an essential step in the synthesis of many important pharmaceutical intermediates. However, the solution to the problem of low biohydroxylation conversion in the biotransformation of DHEA has yet to be found. The effects of natural oils on the course of dihydroxylation of DHEA to 3ß,7α,15α-trihydroxy-5-androsten-17-one (7α,15α-diOH-DHEA) were studied. With rapeseed oil (2 %, v/v) addition, the bioconversion efficiency was improved, and the 7α,15α-diOH-DHEA yield was increased by 40.8 % compared with that of the control at DHEA concentration of 8.0 g/L. Meantime, the ratio of 7α,15α-diOH-DHEA to 7α-OH-DHEA was also increased by 4.5 times in the rapeseed oil-containing system. To explain the mechanism underlying the increase of 7α,15α-diOH-DHEA yield, the effects of rapeseed oil on the pH of the bioconversion system, the cell growth and integrity of Gibberella intermedia CA3-1, as well as the membrane composition were systematically studied. The addition of rapeseed oil enhanced the substrate dispersion and maintained the pH of the system during bioconversion. Cells grew better with favorable integrity. The fatty acid profile of G. intermedia cells revealed that rapeseed oil changed the cell membrane composition and improved cell membrane permeability for lipophilic substrates.
Assuntos
Androstenóis/metabolismo , Desidroepiandrosterona/metabolismo , Gibberella/metabolismo , Óleos de Plantas/química , Biocatálise , Biotransformação , Ácidos Graxos/metabolismo , Ácidos Graxos Monoinsaturados , Gibberella/ultraestrutura , Concentração de Íons de Hidrogênio , Hidroxilação , Óleo de Brassica napusRESUMO
In this study, the isolation of an endophytic fungus from the leaves of the medicinal herb adlay (Coix lacryma-jobi L. var. ma-yuen Stapf) is reported for the first time. The fungus produced Triolein (trioleoylglycerol), a major constituent of triacylglycerols (TAGs) of adlay, in rice medium under shake-flask and bench-scale fermentation conditions. The fungus was identified as Gibberella moniliformis (Fusarium verticillioides) by its morphology and authenticated by ITS analysis (ITS1 and ITS2 regions and the intervening 5.8S rDNA region). Triolein was identified by HPLC-ELSD coupled with APCI-MS and confirmed through comparison with authentic standard. The concentration of triolein produced by G. moniliformis AH13 reached 2.536 ± 0.006 mg/g dry weight of mycelium. Moreover, the EtOAc extract of G. moniliformis AH13 showed strong antitumor activity against four types of tumor cells (A549, HCT116, MDA-MB-231, and SW1990). These results suggest that G. moniliformis AH13 in adlay has significant scientific and industrial potential to meet the pharmaceutical demands and sustainable energy requirements for TAGs in a cost-effective, easily accessible, and reproducible way and is also a potential novel source of natural antitumor bioactive agents.
Assuntos
Antineoplásicos/metabolismo , Coix/microbiologia , Endófitos/classificação , Endófitos/isolamento & purificação , Gibberella/classificação , Gibberella/isolamento & purificação , Trioleína/metabolismo , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Análise por Conglomerados , Meios de Cultura/química , DNA Fúngico/química , DNA Fúngico/genética , DNA Ribossômico/química , DNA Ribossômico/genética , DNA Espaçador Ribossômico/química , DNA Espaçador Ribossômico/genética , Endófitos/genética , Endófitos/metabolismo , Gibberella/genética , Gibberella/metabolismo , Humanos , Espectrometria de Massas , Dados de Sequência Molecular , Filogenia , Folhas de Planta/microbiologia , RNA Ribossômico 5,8S/genética , Análise de Sequência de DNARESUMO
The ascomycete fungus, Fusarium graminearum (teleomorph Gibberella zeae), is the most common causal agent of Fusarium head blight (FHB), a devastating disease for cereal crops worldwide. F. graminearum produces ascospores (sexual spores) and conidia (asexual spores), which can serve as disease inocula of FHB. Meanwhile, Fusarium-infected grains are often contaminated with mycotoxins such as trichothecenes (TRIs), fumonisins, and zearalenones, among which TRIs are related to the pathogenicity of F. graminearum, and these toxins are hazardous to humans and livestock. In recent years, with the complete genome sequencing of F. graminearum, an increasing number of functional genes involved in the production of secondary metabolites, hyphal differentiation, sexual and asexual reproduction, virulence and pathogenicity have been identified from F. graminearum. In this review, the secondary metabolite synthesis, hyphal development and pathogenicity related genes in F. graminearum were thoroughly summarized, and the genes associated with secondary metabolites, sexual reproduction, energy metabolism, and pathogenicity were highlighted.
Assuntos
Fusarium , Genes Fúngicos/genética , Gibberella , Fusarium/genética , Fusarium/metabolismo , Fusarium/patogenicidade , Fusarium/fisiologia , Gibberella/genética , Gibberella/metabolismo , Gibberella/patogenicidade , Gibberella/fisiologia , VirulênciaRESUMO
Gibberelic acid fermentation using extractive methods was carried out in the presence of corn oil and Alamine 336. Gibberella fujikuroi fungus (NRRL 2278) was used to produce gibberellic acid. Oleyl alcohol was a diluting agent for Alamine 336. The effects of oleyl alcohol (100%, v/v), corn oil (5-25%, v/v), the concentration of Alamine 336 in oleyl alcohol, and feeding air were examined in this study. According to the results, oleyl alcohol was not effective on the production. On the other hand, oleyl alcohol solutions containing 15-30% (v/v) Alamine 336 showed effects as a toxic substance. In order to reduce solvent toxicity, corn oil was used. Addition of corn oil increased the concentration of gibberellic acid 1.3-fold compared to the control. Then the effects of immobilization and co-immobilization on extractive gibberelic acid fermentation were investigated. The highest total gibberellic acid concentration of 158.9 mg/L was produced with immobilized cells and feeding air by using extractive fermentation. The yield of gibberellic acid increased about 2.6-fold compared with the shake-flask fermentation (60.5 mg/L) without organic solutions.
Assuntos
Óleo de Milho/farmacologia , Álcoois Graxos/farmacologia , Gibberella/crescimento & desenvolvimento , Giberelinas/biossíntese , Células Imobilizadas/metabolismo , Gibberella/química , Gibberella/metabolismo , Giberelinas/química , Giberelinas/isolamento & purificaçãoRESUMO
Traditionally characterization of microbial proteins is performed by a complex sequence of steps with the final step to be either Edman sequencing or mass spectrometry, which generally takes several weeks or months to be complete. In this work, we proposed a strategy for the characterization of tryptic peptides derived from Giberella zeae (anamorph: Fusarium graminearum) proteins in parallel to intact cell mass spectrometry (ICMS) in which no complicated and time-consuming steps were needed. Experimentally, after a simple washing treatment of the spores, the aliquots of the intact G. zeae macro conidia spores solution, were deposited two times onto one MALDI (matrix-assisted laser desorption ionization) mass spectrometry (MS) target (two spots). One spot was used for ICMS and the second spot was subject to a brief on-target digestion with bead-immobilized or non-immobilized trypsin. Subsequently, one spot was analyzed immediately by MALDI MS in the linear mode (ICMS) whereas the second spot containing the digested material was investigated by MALDI MS in the reflectron mode ("peptide mass fingerprint") followed by protonated peptide selection for MS/MS (post source decay (PSD) fragment ion) analysis. Based on the formed fragment ions of selected tryptic peptides a complete or partial amino acid sequence was generated by manual de novo sequencing. These sequence data were used for homology search for protein identification. Finally four different peptides of varying abundances have been identified successfully allowing the verification that our desorbed/ionized surface compounds were indeed derived from proteins. The presence of three different proteins could be found unambiguously. Interestingly, one of these proteins is belonging to the ribosomal superfamily which indicates that not only surface-associated proteins were digested. This strategy minimized the amount of time and labor required for obtaining deeper information on spore preparations within the nowadays widely used ICMS approach.
Assuntos
Proteínas Fúngicas/metabolismo , Gibberella/metabolismo , Fragmentos de Peptídeos/análise , Esporos Fúngicos/metabolismo , Bases de Dados Genéticas , Proteínas Fúngicas/química , Gibberella/química , Gibberella/classificação , Fragmentos de Peptídeos/genética , Análise de Sequência de Proteína , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Esporos Fúngicos/química , TripsinaRESUMO
Diosgenin is an important raw material in steroid hormone and widely used in pharmaceutical industry. The traditional method for diosgenin production is acidolysis, which causes serious pollution. In order to obtain a cleaner and more efficient approach of diosgenin production, a strain of Gibberella intermedia WX12 (the sexual stage of Fusarium proliferatum) was screened from the strains deposited in our laboratory. This strain converted saponins in Dioscorea zingiberensis C.H. Wright (DZW) into diosgenin. The conversion medium was optimized by statistical experimental design. The optimized conversion medium was as follows (g/L): glucose 20.6, yeast extract 5.0, NaCl 1, K2PO4 3, ZnSO4 x 7H2O 1.5 and saponins 3. Under the optimal conditions, the yield of diosgenin achieved to (31 +/- 0.3) mg/g DZW, which was 3 times higher than that of the original medium.
Assuntos
Dioscorea/química , Diosgenina/metabolismo , Gibberella/metabolismo , Saponinas/metabolismo , Biotransformação , Meios de Cultura , Gibberella/crescimento & desenvolvimentoRESUMO
Three Fusarium species associated with bakanae disease of rice (Fusarium fujikuroi, Fusarium proliferatum, and Fusarium verticillioides) were investigated for their ability to produce fumonisins (FB1 and FB2) under different light conditions, and for pathogenicity. Compared to darkness, the conditions that highly stimulated fumonisin production were yellow and green light in F. verticillioides strains; white and blue light, and light/dark alternation in F. fujikuroi and F. proliferatum strains. In general, all light conditions positively influenced fumonisin production with respect to the dark. Expression of the FUM1 gene, which is necessary for the initiation of fumonisin production, was in accordance with the fumonisin biosynthetic profile. High and low fumonisin-producing F. fujikuroi strains showed typical symptoms of bakanae disease, abundant fumonisin-producing F. verticillioides strains exhibited chlorosis and stunting of rice plants, while fumonisin-producing F. proliferatum strains were asymptomatic on rice. We report that F. fujikuroi might be an abundant fumonisin producer with levels comparable to that of F. verticillioides and F. proliferatum, highlighting the need of deeper mycotoxicological analyses on rice isolates of F. fujikuroi. Our results showed for the first time the influence of light on fumonisin production in isolates of F. fujikuroi, F. proliferatum, and F. verticillioides from rice.