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PubMed2001

Pleureryn, a novel protease from fresh fruiting bodies of the edible mushroom Pleurotus eryngii.

A protease designated pleureryn, with an N-terminal sequence dissimilar from previously reported mushroom metalloendopeptidases and showing only limited resemblance to aspartic proteinases, albeit considerable homology to DNA replication licensing factor, was isolated from fresh fruiting bodies of the edible mushroom Pleurotus eryngii. The purification protocol entailed ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel Blue gel, ion exchange chromatography on CM-Sepharose, and FPLC-gel filtration on Superdex 75. The protease was unadsorbed on DEAE-cellulose but adsorbed on Affi-gel Blue gel and CM-Sepharose. It demonstrated a single band with a molecular weight of 11.5 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Pleureryn demonstrated a protease activity of 9364 U/mg toward casein. It exhibited a pH optimum of 5.0 and a temperature optimum of 45 degrees C, with substantial activity remaining at high temperatures and pH 4 and 12. The activity of the protease was adversely affected by pepstatin A, indicating that it is an aspartic protease. PMSF, trypsin inhibitor, and EDTA exerted no striking effect, suggesting that it is neither a serine protease nor a metalloprotease. It inhibited translation in a rabbit reticulocyte lysate system with an IC(50) of 20 nM. Pleureryn also exhibited some inhibitory activity against HIV-1 reverse transcriptase, reminiscent of a suppressive action of HIV-1 protease on its homologous reverse transcriptase but was devoid of ribonuclease, deoxyribonuclease, and antifungal activities.

Amino Acid Sequence↗

PubMed2001

Antioxidant and immunostimulating activities of the fruiting bodies of Paecilomyces japonica, a new type of Cordyceps sp.

Cordyceps is negative for its many biological activities and a tonic for restoring vital functions in traditional Chinese medicine. In an effort to evaluate the pharmacological effects, including the antiaging effect of the fruiting bodies of the cultivated Paecilomyces japonica fungus, a new type of Cordyceps sp. was investigated. This investigation was focused on ultimately revealing its biologically active principles, its effects on free-radical scavenging enzymes, lipid peroxidation, as well as its immunological functions. As a result, both water and methanol extracts were found to cause not only significant increases in rat liver cytosolic SOD, catalase, and GSEH-px activities, but also a significant decrease in MDA production in TBA reactant assay in rats. The extracts also showed immunostimulating activity as measured by carbon clearance, weight-loaded forced swimming performances, and immobilizing stress in mice. Using bioassay-guided systematic fractionation of the extracts, two pure compounds were isolated as active principles from low molecular-weight fraction, a protein-bound polysaccharide was isolated that showed a marked increase in the liver enzyme activities, as well as a significant inhibition of lipid peroxidation.

Adjuvants, Immunologic↗

PubMed2006

Apoptotic effects of Antrodia cinnamomea fruiting bodies extract are mediated through calcium and calpain-dependent pathways in Hep 3B cells.

Antrodia cinnamomea is well known in Taiwan as a traditional medicine for treating cancer and inflammation. The purpose of this study was to evaluate the apoptotic effects of ethylacetate extract from A. cinnamomea (EAC) fruiting bodies in Hep 3B, a liver cancer cell line. EAC decreased cell proliferation of Hep 3B cells by inducing apoptotic cell death. EAC treatment increased the level of calcium (Ca2+) in the cytoplasm and triggered the subsequent activation of calpain and caspase-12. EAC also initiated the mitochondrial apoptotic pathway through regulation of Bcl-2 family proteins expression, release of cytochrome c, and activation of caspase-9 in Hep 3B cells. Furthermore, the mitochondrial apoptotic pathway amplified the calpain pathway by Bid and Bax interaction and Ca2+ translocation. We have therefore concluded that the molecular mechanisms during EAC-mediated proliferation inhibition in Hep 3B cells were due to: (1) apoptosis induction, (2) triggering of Ca2+/calpain pathway, (3) disruption of mitochondrial function, and (4) apoptotic signaling being amplified by cross-talk between the calpain/Bid/Bax and Ca2+/mitochondrial apoptotic pathways.

Apoptosis↗

PubMed1992

Polysaccharides in fungi. XXIX. Structural features of two antitumor polysaccharides from the fruiting bodies of Armillariella tabescens.

An antitumor polysaccharide containing peptide moieties AT-HW ([alpha]D + 31 degrees in water) and an antitumor polysaccharide AT-AL ([alpha]D + 209 degrees in 1 M sodium hydroxide) were isolated from hot-water extract and the alkaline extract of the fruiting bodies of Armillariella tabescens, respectively. Chemical structures of AT-HW and AT-AL were investigated by a combination of chemical and spectroscopic methods. The results indicate that the major constituent of AT-HW (molecular weight, 105000), a heteroglycan, is composed primarily of beta-(1----6)-linked D-glucopyranosyl and D-galactopyranosyl residues, and contains their branched residues and terminal sugar (gluco-, manno-, and fucopyranose) residues, in addition to beta-(1----3)-linked D-glucopyranosyl residues, while AT-AL (molecular weight, 93000) is chiefly composed of alpha-(1----3)-linked D-glucopyranosyl residues.

Antineoplastic Agents↗

PubMed2001

Two new steroidal derivatives from the fruit body of Chlorophyllum molybdites.

Two new steroid derivatives, (22E,24R)-3alpha-ureido-ergosta-4,6,8(14),22-tetraene (1) and (22E,24R)-5alpha,8alpha-epidioxyergosta-6,9,22-triene-3beta-ol 3-O-beta-D-glucopyranoside (2) were isolated from the fruit bodies of Chlorophyllum molybdites (Agaricaceae). The structures were established by spectroscopic and chemical methods. These compounds exhibited cytotoxicity against Kato III cells.

Agaricales↗

PubMed2001

Laboratory culture of the myxomycetes: formation of fruiting bodies of Didymium bahiense and its plasmodial production of Makaluvamine A.

Laboratory cultures of more than 100 strains of myxomycetes were investigated, and the spore germinations of six strains were observed. The plasmodium of the myxomycete Didymium bahiense was cultured on oatmeal agar plates in a laboratory. The formation of fruiting bodies was observed in a plate culture. From the cultured organisms, a marine sponge metabolite, makaluvamine A (1), was isolated and identified on the basis of spectral data.

Animals↗

PubMed1999

Isolation of a new ribonuclease from fresh fruiting bodies of the straw mushroom.

Although ribonucleases have been isolated from numerous organisms in the animal and plant kingdoms, only a few have been reported from mushrooms. In this investigation, a novel ribonuclease, with an N-terminal amino acid sequence distinctly different from those of the previously published mushroom ribonucleases, was isolated from the fruiting bodies of the straw mushroom. It exhibited a molecular weight of 42.5 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis which was larger than those of ribonucleases from other organisms. The isolation procedure entailed extraction with aqueous buffer, (NH(4))(2)SO(4) precipitation, ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, ion exchange chromatography on CM-Sepharose CL6B, and gel filtration on Superdex 75 using fast protein liquid chromatography. Straw mushroom ribonuclease was relatively specific for polyU and hydrolyzed yeast tRNA with a pH optimum between 6.5 and 7.5.

Amino Acid Sequence↗

PubMed2000

Structures of new ceramides from the fruit bodies of Grifola frondosa.

Four new phytosphingosine-type ceramides, (2S,3S,4R)-2-[(2'R)-2'-hydroxydocosanoylamino]-1,3,4-octadecane triol (1), (2S,3S,4R)-2-[(2'R)-2'-hydroxytricosanoylamino]-1,3,4-octadecan etriol (2), (2S,3S,4R)-2-[(2'R)-2'-hydroxypentacosanoylamino]-1,3,4-octadec anetriol (3) and (2S,3S,4R)-2-[(2'R)-2'-hydroxyhexacosanoylamino]-1,3,4-octadeca netriol (4), have been isolated from the fruit bodies of Grifola frondosa. The structures of the new compounds were elucidated on the basis of their spectral data.

Ceramides↗

PubMed2006

Isolation of protoplast from aerial mycelia and fruit-body formation from regenerated mycelia in Pleurotus cornucopiae.

We have developed a simple and efficient method for protoplast isolation from Pleurotus cornucopiae. Protoplasts were isolated from aerial mycelia cultured on potato dextrose agar medium without time-consuming propagation in liquid culture. Protoplast yield was significantly increased by means of a decompressing pretreatment of mycelia in enzyme solution and a subsequent enzyme reaction with vibrational mixing. The isolated protoplasts regenerated mycelia and these mycelia formed fruit-bodies without any morphological abnormalities.

Pleurotus↗

PubMed2003

Global mutational analysis of NtrC-like activators in Myxococcus xanthus: identifying activator mutants defective for motility and fruiting body development.

The multicellular developmental cycle of Myxococcus xanthus requires large-scale changes in gene transcription, and recent findings indicate that NtrC-like activators play a prominent role in regulating these changes. In this study, we made insertions in 28 uncharacterized ntrC-like activator (nla) genes and found that eight of these insertions cause developmental defects. Hence, these results are consistent with the idea that M. xanthus uses a series of different NtrC-like activators during fruiting body development. Four of the eight developmental mutants we identified have motility defects. The nla1, nla19, and nla23 mutants show S-motility defects, while the nla24 mutant shows defects in both S-motility and A-motility. During development, aggregation of the nla1, nla19, and nla23 mutants is delayed slightly and the nla24 mutant shows no signs of aggregation or sporulation. The nla4, nla6, nla18, and nla28 mutants have no appreciable loss in motility, but they fail to aggregate and to sporulate normally. The nla18 mutant belongs to a special class of developmental mutants whose defects can be rescued when they are codeveloped with wild-type cells, suggesting that nla18 fails to produce a cell-cell signal required for development. The three remaining activator mutants, nla4, nla6, and nla28, appear to have complex developmental phenotypes that include deficiencies in cell-cell developmental signals.

Bacterial Proteins↗

PubMed2006

Inhibition of anaphylaxis-like reaction and mast cell activation by water extract from the fruiting body of Phellinus linteus.

Mast cell-mediated anaphylactic reaction is involved in many allergic diseases such as asthma and allergic rhinitis. Phellinus linteus has been used as a traditional herb medicine in oriental countries and is known to have anti-tumor, immunomodulatory, anti-inflammatory, and anti-allergic activities. However, roles of Phellinus linteus in the mast cell-mediated anaphylactic reactions have not fully been examined. In the present study, we have investigated the effects of water extract from the fruiting body of Phellinus linteus (WEPL) on mast cell-mediated anaphylaxis-like reactions. Oral administration of WEPL inhibited the compound 48/80-induced systemic anaphylaxis-like reaction and ear swelling response. WEPL also inhibited the anti-dinitrophenyl (DNP) IgE-mediated passive systemic and cutaneous anaphylaxis. WEPL had no cytotoxicity on rat peritoneal mast cells (RPMC). WEPL dose-dependently reduced histamine release from RPMC activated by compound 48/80 or anti-DNP IgE. Moreover, WEPL decreased the compound 48/80-induced calcium uptake into RPMC. Furthermore, WEPL increased the level of intracellular cAMP and significantly inhibited the compound 48/80-induced cAMP reduction in RPMC. These results suggest that WEPL may serve as an effective therapeutic agent for allergic diseases.

Anaphylaxis↗

PubMed1999

New lanostanoid glycosides from the fruit body of laetiporus versisporus

Four new lanostanoid glycosides, 3beta, 7alpha-dihydroxy-24-methylene-lanost-8-en-21-oic acid 7-O-beta-D-glucopyranoside (1), 3beta,7alpha-dihydroxy-lanost-8, 24-dien-21-oic acid 7-O-beta-D-glucopyranoside (2), 7alpha-hydroxy-3-keto-lanost-8,24-dien-21-oic acid 7-O-beta-D-glucopyranoside (3), and 7alpha-hydroxy-3-keto-24-methylene-lanost-8-en-21-oic acid 7-O-beta-D-glucopyranoside (4) were isolated from the fruit bodies of Laetiporus versisporus. Compounds 1-4 were named laetiposides A-D, respectively. Their structures were established by extensive NMR experiments and chemical methods.

Journal Article↗

PubMed2001

eEF1A Controls ascospore differentiation through elevated accuracy, but controls longevity and fruiting body formation through another mechanism in Podospora anserina.

Antisuppressor mutations in the eEF1A gene of Podospora anserina were previously shown to impair ascospore formation, to drastically increase life span, and to permit the development of the Crippled Growth degenerative process. Here, we show that eEF1A controls ascospore formation through accuracy level maintenance. Examination of antisuppressor mutant perithecia reveals two main cytological defects, mislocalization of spindle and nuclei and nuclear death. Antisuppression levels are shown to be highly dependent upon both the mutation site and the suppressor used, precluding any correlation between antisuppression efficiency and severity of the sporulation impairment. Nevertheless, severity of ascospore differentiation defect is correlated with resistance to paromomycin. We also show that eEF1A controls fruiting body formation and longevity through a mechanism(s) different from accuracy control. In vivo, GFP tagging of the protein in a way that partly retains its function confirmed earlier cytological observation; i.e., this factor is mainly diffuse within the cytosol, but may transiently accumulate within nuclei or in defined regions of the cytoplasm. These data emphasize the fact that the translation apparatus exerts a global regulatory control over cell physiology and that eEF1A is one of the key factors involved in this monitoring.

Alleles↗

PubMed1991

Hydrophobin Genes Involved in Formation of Aerial Hyphae and Fruit Bodies in Schizophyllum.

Fungi typically grow by apical extension of hyphae that penetrate moist substrates. After establishing a branched feeding mycelium, the hyphae differentiate and grow away from the substrate into the air where they form various structures such as aerial hyphae and mushrooms. In the basidiomycete species Schizophyllum commune, we previously identified a family of homologous genes that code for small cysteine-rich hydrophobic proteins. We now report that the encoded hydrophobins are excreted in abundance into the culture medium by submerged feeding hyphae but form highly insoluble complexes in the walls of emerging hyphae. The Sc3 gene encodes a hydrophobin present in walls of aerial hyphae. The homologous Sc1 and Sc4 genes, which are regulated by the mating-type genes, encode hydrophobins present in walls of fruit body hyphae. The hydrophobins are probably instrumental in the emergence of these aerial structures.

Journal Article↗

PubMed2006

3-Hydroxy-3-methylglutaryl-coenzyme A (CoA) synthase is involved in biosynthesis of isovaleryl-CoA in the myxobacterium Myxococcus xanthus during fruiting body formation.

Isovaleryl-coenzyme A (IV-CoA) is the starting unit for some secondary metabolites and iso-odd fatty acids in several bacteria. According to textbook biochemistry, IV-CoA is derived from leucine degradation, but recently an alternative pathway that branches from the well-known mevalonate-dependent isoprenoid biosynthesis has been described for myxobacteria. A double mutant was constructed in Myxococcus xanthus by deletion of genes involved in leucine degradation and disruption of mvaS encoding the 3-hydroxy-3-methylglutaryl-coenzyme A synthase. A dramatic decrease of IV-CoA-derived iso-odd fatty acids was observed for the mutant, confirming mvaS to be involved in the alternative pathway. Additional quantitative real-time reverse transcription-PCR experiments indicated that mvaS is transcriptionally regulated by isovalerate. Furthermore, feeding studies employing an intermediate specific for the alternative pathway revealed that this pathway is induced during fruiting body formation, which presumably increases the amount of IV-CoA available when leucine is limited.

Acyl Coenzyme A↗

PubMed2004

Molecular cloning of a new laccase from the edible straw mushroom Volvariella volvacea: possible involvement in fruit body development.

Cloning of a laccase-encoding cDNA from the edible straw mushroom, Volvariella volvacea, was performed using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends. The cDNA of the putative laccase gene (lac4) consisted of 1689 bp, including an open reading frame encoding a 23-amino acid signal peptide at the N-terminal end and a 540-amino acid mature protein with a predicted molecular mass of 58173 Da and a pI value of 6.1. The 10 histidine residues and one cysteine residue required to co-ordinate the four copper atoms at the active site of the protein were all conserved. The amino acid sequence of V. volvacea lac4 has a high degree of identity with other basidiomycete laccases. Transcription of the laccase gene was analysed by RT-PCR and, unlike many other laccase genes, shown to be regulated independently of either copper or aromatic compounds under the test conditions. However, the laccase gene is strongly expressed during that part of the mushroom developmental cycle involving fruit body morphogenesis.

Agaricales↗

PubMed2002

New lanostanoids, elfvingic acids A-H, from the fruit body of Elfvingia applanata.

Eight new lanostanoids, 12alpha,15beta-dihydroxy-3,7,11,23-tetraoxolanost-8,(20Z)(22)-dien-26-oic acid (1), 7beta,8beta-epoxy-15beta,20S-dihydroxy-3,12,23-trioxolanost-9(11),16-dien-26-oic acid (2), 7beta,8beta-epoxy-3beta,15beta,20S-trihydroxy-12,23-dioxolanost-9(11),16-dien-26-oic acid (3), 7beta,8beta-epoxy-2alpha,3beta,15beta,20S-tetrahydroxy-12,23-dioxolanost-9(11),16-dien-26-oic acid (4), 7beta,8beta-epoxy-3beta,15beta,20S,28-tetrahydroxy-12,23-dioxolanost-9(11),16-dien-26-oic acid (5), 7beta,8beta-epoxy-3beta,15beta,19,20S-tetrahydroxy-12,23-dioxolanost-9(11),16-dien-26-oic acid (6), 8beta,15beta,20S-trihydroxy-3,7,12,23-tetraoxolanost-9(11),16-dien-26-oic acid (7), 7alpha,8alpha-epoxy-15beta,23xi-dihydroxy-12-oxo-3,4-secolanost-4(28),9(11),(20Z)(22)-trien-3,26-dioic acid (8), and the methyl ester of 8 (8a) were isolated from the fruit bodies of Elfvingia applanata. Their structures were established primarily by NMR experiments, and their biological activity against Kato III and Ehlrich cells was investigated.

Antineoplastic Agents, Phytogenic↗

PubMed1996

Identification and characterization of FrzZ, a novel response regulator necessary for swarming and fruiting-body formation in Myxococcus xanthus.

The frz genes of Myxococcus xanthus constitute a signal-transduction pathway that processes chemotactic information in a manner analogous to that found in enteric bacteria. Ultimately, these genes regulate the frequency of individual cell reversal. We report here the identification of a novel component of this signal-transduction pathway, designated frzZ, which was discovered as an open reading frame located 5' to the frz operon but transcribed in the opposite orientation. The translational start site of frzZ is 170 base pairs from that of frzA.frzZ utilizes a promoter similar to the sigma 70 promoters of Escherichia coli, and encodes a 290-amino-acid soluble protein, FrzZ (M(r) 30,500). FrzZ contains two domains, both of which show strong homology to CheY and other members of the response-regulator family. Linking these domains is a 39-amino-acid region that is very rich in alanine and proline (38% Ala and 33% Pro). A frzZ null mutant showed abnormally low reversal rates when compared to the wild-type control and was unable to form fruiting bodies on starvation medium, but it did form 'frizzy' aggregates. In addition, the frzZ mutant was defective in swarming, particularly on soft agar (0.3% w/v). However, unlike most frz mutants, the frzZ mutant was able to respond to attractants and repellents in the spatial chemotaxis assay. The discovery of FrzZ demonstrates that the M. xanthus frz signal-transduction pathway utilizes multiple response-regulator (CheY-like) proteins.

Amino Acid Sequence↗