Chemotaxonomy of Gibberella zeae with special reference to production of trichothecenes and zearalenone.

By adopting a single-spore isolation technique, 113 isolates of Gibberella zeae, the perfect stage of Fusarium graminearum, were isolated from rice stubbles in barley and wheat fields and tested for production of trichothecenes and zearalenone on rice grains. Of the isolates, 93% produced the trichothecenes, and they could be subdivided into two chemotaxonomic groups: nivalenol and fusarenon-X producers and deoxynivalenol and 3-acetyldeoxynivalenol producers. No cross production of these two types of trichothecenes was observed in these isolates. Zearalenone was detected in 68% of the isolates, but no clear relationship could be observed regarding its position with respect to the two chemotaxonomic groups.     

Expression in Escherichia coli of chemically synthesized gene for the human immune interferon.

A 454 base pair fragment of double stranded DNA consisting of a gene for a human immune interferon (hIFN-gamma), initiation and termination signals plus appropriate restriction endonuclease sites, was totally synthesized. The synthesis involved preparation of 62 oligodeoxyribonucleotides by rapid, solid phase procedures, and enzymatic ligation of the oligonucleotides. This synthetic gene was expressed in E. coli under the control of the lac UV5 promoter. The product has antiviral activity which was acid labile and completely neutralized by antiserum to hIFN-gamma but not by antiserum to hIFN-alpha or hIFN-beta. Molecular weight of hIFN-gamma produced by E. coli was estimated to be about 32,000 and 17,000 by gel filtration and SDS-polyacrylamide gel electrophoresis respectively.     

Enzymic conversion of 11,12-leukotriene A4 to 11,12-dihydroxy-5,14-cis-7,9-trans-eicosatetraenoic acid. Purification of an epoxide hydrolase from the guinea pig liver cytosol

(11S,12S)-Epoxy-5,14-cis-7,9-trans-eicosatetraenoic acid (11,12-leukotriene A4) was nonenzymically converted to seven compounds: two diastereomers of (12S)-hydroxyeicosatetraeno-delta-lactones (major products), two diastereomers of (5,12S)-dihydroxyeicosatetraenoic acid and three stereoisomers of (11,12S)-dihydroxyeicosatetraenoic acid. Among these compounds, (11R,12S)-dihydroxy-5,14-cis-7,9-trans-eicosatetraenoic acid proved to be the only enzymic product. This hydrolysis activity was present in the cytosol fractions of various tissues of guinea pig such as liver, adrenal gland, small intestine, and brain. We purified the epoxide hydrolase to an apparent homogeneity from the guinea pig liver. The enzyme had a molecular weight of 60,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and an isoelectric point of 7.3. The partial amino acid sequence was different from that of the microsomal enzyme. Km and Vmax values for 11,12-leukotriene A4 were 18 microM and 2.4 mumol/min/mg protein, respectively. These results indicate that 11,12-dihydroxyeicosatetraenoic acid is enzymically synthesized from 11,12-leukotriene A4 by the action of the cytosolic epoxide hydrolase in vitro.     

Enhancement of the Division of Equisetum arvense Protoplasts in Culture by Activated Charcoal and Their Further Development

Protoplasts were isolated from subcultured gametophytes of Equisetumarvense by treatment with Driselase and then cultured in vitro.Addition of activated charcoal (AC) to the culture medium enhancedthe rate of cell division, as well as the survival of both protoplastsand regenerated protoplasts. However, subsequent division ofcells was not observed after one or two cycles of replicationin cultures supplemented with AC. When regenerated protoplastswere transferred to fresh medium without AC 3 to 5 weeks afterthe first plating, the transferred cells formed rhizoids anddeveloped into small, young gametophytes without the prior formationof cell clusters or calluses. Furthermore, sprophytic shootsdifferentiated from the protoplast-derived gametophytes whenthey were cultured on medium supplemented with 6-benzylaminopurine(BA). (Received April 5, 1990; Accepted July 30, 1990)     

Interleukin-4 enhances in vitro mineralization in human osteoblast-like cells.

The effect of human interleukin-4 (hIL-4) on mineralization in human osteoblast-like cells was investigated. Confluent cells were incubated with hIL-4 for 16 or 30 days in the presence or absence, respectively, of alpha-glycerophosphate (alpha-GP), which accelerates the mineralization process. hIL-4 (0.3 ng/ml) induced mineralization with 1.9-, 26- and 37-fold increases of hydroxyproline, calcium, and osteocalcin content, respectively, in the presence of alpha-GP. Mineralization was not induced with other cytokines, hIL-1, hIL-2, hIL-6, or mIL-4. hIL-4 also induced mineralization in the absence of alpha-GP in a manner different from that of 1 alpha, 25(OH)2 vitaminD3 (1,25(OH)2VD3). These findings suggest that IL-4 may play an important role in bone formation.     

A 1H-NMR study of the solution conformation of cyclo(GRGDSPA): conformational effects on the physiological activity.

Solution conformations of cyclo(GRGDSPA) have been analyzed by the use of two-dimensional proton nuclear magnetic resonance spectroscopy and the dynamical simulated annealing calculation. It has been shown that the RGDS segment in cyclo(GRGDSPA) takes a beta-turn conformation. We have concluded that this beta-turn conformation is essential for the physiological activity of cyclo(GRGDSPA).     

Analysis of chorion hardening of eggs of rainbow trout, Oncorhynchus mykiss

We estimated changes of chorion hardness of rainbow trout (Oncorhynchus mykiss) egg by the use of three parameters, namely increase of resistance of an egg to rupture by extraneously applied pressure, decrease of solubility of chorion proteins in 8 mol/L urea and a change in the content of γ-glutamyl-ε-lysine crosslink. Unfertilized egg chorions became hardened after egg activation. During chorion hardening, 49, 56 and 65 kDa protein components of the chorion gradually disappeared, high molecular weight intermediates (113,160–170 and higher than 250 kDa) were newly formed and, finally, all components became undetectable by sodium dodecylsulfate-polyacrylamide gel electrophoresis. The content of γ-glutamyl-ε-lysine (γ-Glu-ε-Lys) crosslink in the chorion increased after hardening. Chorion hardening was inhibited by the incorporation of monodansyl-cadaverine, a competitive inhibitor for transglutaminase (TGase), into the chorions. TGase activity was detected in unfertilized eggs and localized in the chorion fraction rather than in the ooplasmic fraction. The findings suggest that chorion hardening depends upon polymerization of the chorion components by TGase-dependent γ-Glu-ε-Lys crosslink formation.