Ketoisophorone transformation by Marchantia polymorpha and Nicotiana tabacum cultured cells

Stereospecific olefin (C=C) and carbonyl (C=O) reduction of the readily available prochiral compound ketoisophorone (2,2,6-trimethyl-2-cyclohexene-1,4-dione) (1) by Marchantia polymorpha and Nicotiana tabacum cell suspension cultures produce the chiral products (6R)-levodione (2), (4R,5S)-4-hydroxy-3,3,5-trimethylcyclohexanone (3), and (4R,6R)-actinol (4) as well as the minor components (4R)-hydroxyisophorone (5) and (4S)-phorenol (6).     

Characterization of cryptic flagellin genes in Shigella boydii and Shigella dysenteriae.

Flagellin (fliC) genes of 12 Shigella boydii and five Shigella dysenteriae strains were characterized. Though these strains are nonmotile, the cryptic fliCSB gene, cloned from S. boydii strain C3, is functional for expression of flagellin. It consists of 1,704 bp, and encodes 568 amino acid residues (57,918 Da). The fliCSD gene from S. dysenteriae strain 16 consists of 1,650 bp encoding 549 amino acid residues (57,591 Da) and contains an IS1 element inserted in its 3' end. The two genes are composed of the 5'-constant, central variable and 3'-constant sequences, like other known fliC genes. The two genes share high homology in nucleotide and amino acid sequences with each other and also with the Escherichia coli fliCE gene, indicating that both genes are closely related to the fliCE gene. Comparison of the central variable sequences of six different fliC genes showed that the fliCSB and fliCSD genes share low homology in amino acid sequence with the other fliC genes, suggesting that they encode antigenic determinants intrinsic to respective subgroups. However, Southern blotting using as probes the central variable sequences of several fliC genes showed that four of 12 S. boydii strains have a fliC gene similar to that of Shigella flexneri, and that among five fliC genes from S. dysenteriae strains, one is similar to that of S. flexneri, two are similar to that of S. boydii, and only one is unique to S. dysenteriae. Some of these variant alleles were verified by immunoblotting with flagellins produced from cloned fliC genes. The presence of variant fliC alleles in S. boydii and S. dysenteriae indicates that subdivision into subgroups does not reflect the ancestral flagella H antigenic relationships. These data will be useful in considering the evolutionary divergence of the Shigella spp..     

Inhibition of growth and aflatoxin biosynthesis of Aspergillus flavus by extracts of some Egyptian plants

The effect of five different concentrations (2, 4, 6, 8 and 10 mg ml-1) of an aqueous extracts of Lupinus albus, Ammi visnaga and Xanthium pungens were tested on growth and aflatoxin production by Aspergillus flavus in a chemically defined medium. All the plants inhibited mycelial growth and aflatoxin formation. The inhibitory effect was proportional with the applied concentration. Growth and aflatoxin production appeared to be correlated processes. The nature of the plant extract also affected the ratio of B1 to B2, and there was no correlation between the inhibition of aflatoxins or growth of the fungus and the resultant B1: B2 ratio.     

Isolation of alpha-lactalbumin,beta-lactoglobulin,and bovine serum albumin from cow's milk using gel filtration and anion-exchange chromatography including evaluation of their antigenicity

The aim of this study was to introduce a simple, reproducible, and less expensive method for isolation of alpha-lactalbumin, beta-lactoglobulin, and bovine serum albumin from cow's milk while retaining their antigenicity. Whey (lactoserum) was obtained by isolating casein from defatted milk using hydrochloric acid. Globulins were then precipitated from whey by half-saturated ammonium sulfate and beta-lactoglobulin was purified further using Sephadex G-50 gel filtration. The proteins in the supernatant were also fractionated using diethylaminoethyl cellulose chromatography in which beta-lactoglobulin was separated from alpha-lactalbumin and bovine serum albumin. The latter two proteins that co-eluted in anion-exchange chromatography were then gently isolated from each other by Sephadex G-50 gel filtration. Pure beta-lactoglobulin was also obtained by anion-exchange chromatography of the ammonium sulfate-precipitated globulins. Using enzyme-linked immunosorbent assay (ELISA), Western blotting, and ELISA inhibition assay, antigenicity of the purified proteins was evaluated. Our results showed high purity and well-preserved antigenicity of alpha-lactalbumin, beta-lactoglobulin, and bovine serum albumin thus purified.     

The effect of endomorphins on the release of 3H-norepinephrine from rat nucleus tractus solitarii slices

We used two, 3-min field stimulation cycles 30 min apart (S1, S2) in 3H-norepinephrine-loaded, superfused rat nucleus tractus solitarii-dorsal motor vagal nucleus (NTS-DVN) slices. The stimulation-induced release was expressed as the area above the baseline. Drugs were introduced 12 min before S2 and drug actions were characterized in terms of alterations of S2/S1 ratios. The S2/S1 ratio was 1.047 (0.946-1.159, n = 4, geometric mean and 95% confidence interval) in controls and 0.336 (0.230-0.490, n = 3), 0.726 (0.590-0.892, n = 4), 0.613 (0.594-0.683, n = 4) and 0.665 (0.500-0.886, n = 4) in the presence of 10(-6) M clonidine, D-Ala(2),MePhe(4),Gly(5)-ol-enkephalin (DAMGO), endomorphin-1 (Tyr-Pro-Trp-Phe-NH(2), EM-1) and -2 (Tyr-Pro-Phe-Phe-NH(2), EM-2) [the latter two in the presence of 10(-4) M diprotin A, an inhibitor of dipeptidyl-aminopeptidase IV (DAP-IV) enzyme]. The effect of DAMGO at 10(-5) M was significantly higher than at 10(-6) M, whereas the effect of endomorphins did not differ at the two concentration levels. Diprotin A potentiated only very modestly the action of endomorphins. These data (a) confirm the presence of functional mu-opioid receptors in the vagal complex, (b) render it likely that the enzymic degradation of endomorphins is not a highly effective process in brain slices and (c) may suggest that the apparent ceiling in the effect of endomorphins might be related to their partial agonist property.     

An Arabidopsis pex10 null mutant is embryo lethal, implicating peroxisomes in an essential role during plant embryogenesis

Peroxisomes participate in many important functions in plants, including seed reserve mobilization, photorespiration, defense against oxidative stress, and auxin and jasmonate signaling. In mammals, defects in peroxisome biogenesis result in multiple system abnormalities, severe developmental delay, and death, whereas in unicellular yeasts, peroxisomes are dispensable unless required for growth of specific substrates. PEX10 encodes an integral membrane protein required for peroxisome biogenesis in mammals and yeast. To investigate the importance of PEX10 in plants, we characterized a Ds insertion mutant in the PEX10 gene of Arabidopsis (AtPEX10). Heterozygous AtPEX10::dissociation element mutants show normal vegetative phenotypes under optimal growth conditions, but produce about 20% abnormal seeds. The embryos in the abnormal seeds are predominantly homozygous for the disruption allele. They show retarded development and some morphological abnormalities. No viable homozygous mutant plants were obtained. AtPEX10 fused to yellow fluorescent protein colocalized with green fluorescent protein-serine-lysine-leucine, a well-documented peroxisomal marker, suggesting that AtPEX10 encodes a peroxisomal protein that is essential for normal embryo development and viability.     

Common loci underlie field resistance to soybean sudden death syndrome in Forrest,Pyramid, Essex,and Douglas

Soybean [Glycine max (L.) Merr.] sudden death syndrome (SDS) caused by Fusarium solani f. sp. glycines results in severe yield losses. Resistant cultivars offer the most-effective protection against yield losses but resistant cultivars such as ’Forrest’ and ’Pyramid’ vary in the nature of their response to SDS. Loci underlying SDS resistance in ’Essex’ × Forrest are well defined. Our objectives were to identify and characterize loci and alleles that underlie field resistance to SDS in Pyramid×’Douglas’. SDS disease incidence and disease severity were determined in replicated field trials in six environments over 4 years. One hundred and twelve polymorphic DNA markers were compared with SDS disease response among 90 recombinant inbred lines from the cross Pyramid×Douglas. Two quantitative trait loci (QTLs) for resistance to SDS derived their beneficial alleles from Pyramid, identified on linkage group G by BARC-Satt163 (261-bp allele, P=0.0005, R²=16.0%) and linkage group N by BARC-Satt080 (230-bp allele, P=0.0009, R²=15.6%). Beneficial alleles of both QTLs were previously identified in Forrest. A QTL for re- sistance to SDS on linkage group C2 identified by BARC-Satt307 (292-bp allele, P=0.0008, R²=13.6%) derived the beneficial allele from Douglas. A beneficial allele of this QTL was previously identified in Essex. Recombinant inbred lines that carry the beneficial alleles for all three QTLs for resistance to SDS were significantly (P≤0.05) more resistant than other recombinant inbred lines . Among these recombinant inbred lines resistance to SDS was environmentally stable. Therefore, gene pyramiding will be an effective method for developing cultivars with stable resistance to SDS. Received: 20 October 1999 / Accepted: 22 May 2001     

Anti-viral activity of red microalgal polysaccharides against retroviruses

Red microalgal polysaccharides significantly inhibited the production of retroviruses (murine leukemia virus- MuLV) and cell transformation by murine sarcoma virus(MuSV-124) in cell culture. The most effective inhibitory effect of these polysaccharides against both cell transformation and virus production was obtained when the polysaccharide was added 2 h before or at the time of infection. Although, addition of the polysaccharide post-infection significantly reduced the number of transformed cells, but its effect was less marked than that obtained when the polysaccharide was added before or at the time of infection.The finding that the inhibition of cell transformation by MuSV-124 was reversible after removal of the polysaccharide suggested that microalgal polysaccharides inhibited a late step after provirus integration into the host genome. In conclusion, our findings could support the possibility that the polysaccharide may affect early steps in the virus replication cycle, such as virus absorption into the host cells, in addition to its effect on a late step after provirus integration.     

Mimicking a cytoskeleton by coupling poly(N-isopropylacrylamide) to the inner leaflet of liposomal membranes: effects of photopolymerization on vesicle shape and polymer architecture

Networks of N-isopropylacrylamide (NIPAM) copolymers, coupled to spherical phospholipid bilayers, are suitable as a model for the study of the interaction between the cytoskeleton and cellular membranes, as well as for promising new drug delivery systems with triggerable drug release properties and improved stability. In this article, we describe a simple preparation technique for liposomes from egg phosphatidyl choline (EPC) encapsulating a cross-linked NIPAMminus signTEGDM copolymer skeleton (tetraethylene glycol dimethacrylate, TEGDM) which is coupled only to the inner monolayer by a novel membrane anchor monomer. Polymerization in the lipid vesicles was initiated at the inner membrane surface by the radical initiator 2,2-diethoxy-acetophenone (DEAP) permeating through the membrane from the outside. The effects of photopolymerization and polymer formation on vesicle shape and membrane integrity were studied by transmission electron microscopy (TEM), cryo-TEM, and atomic force microscopy (AFM). Upon UV irradiation, approximately 100% of the vesicles contained a polymer gel and only occasional changes in the spherical shape of the liposomes were observed. The architecture of the polymer network inside the liposomal compartment was determined by the conditions of the photopolymerization. Composite structures of polymer hollow spheres or solid spheres, respectively, tethered to spherical membrane vesicles were produced. The increased stability of the polymer-tethered lipid bilayers against solubilization by sodium cholate, compared to pure EPC vesicles, was determined by radiolabeling the lipid membrane.