Protein glycosylation.

Glycan structures can modulate the biological properties and functions of glycoproteins. This has been shown by investigation of the biological activities and glycan structures of several recombinant glycoproteins. Glycan structures of glycoproteins differ according to the species and tissue producing them, and selection of an appropriate host-cell type can generate recombinant glycoproteins with new characteristics.     

Purification and characterization of eugenol dehydrogenase from Pseudomonas fluorescens E118

Pseudomonas fluorescens E118 was isolated from soil as an effective eugenol-degrading organism by a screening using eugenol as enrichment substrate. The first enzyme involved in the degradation of eugenol in this organism, eugenol dehydrogenase, was purified after induction by eugenol, and the purity of the enzyme was shown by SDS-PAGE and gel-permeation HLPC. The enzyme is a heterodimer that consists of a 10-kDa cytochrome c and a 58-kDa subunit. The larger subunit presumably contains flavin, suggesting a flavocytochrome c structure and an electron transfer via flavin and cytochrome c during dehydrogenation. The activity of the purified enzyme depended on the addition of a final electron acceptor such as phenazine methosulfate, 2,6-dichlorophenol-indophenol, cytochrome c, or potassium ferricyanide. The enzyme catalyzed the dehydrogenation of three different 4-hydroxybenzylic structures including the conversion of eugenol to coniferyl alcohol, 4-alkylphenols to 1-(4-hydroxyphenyl)alcohols, and 4-hydroxybenzylalcohols to the corresponding aldehydes. The catalytic and structural similarity between this enzyme and a Penicillium vanillyl-alcohol oxidase and 4-alkylphenol methylhydroxylases from several Pseudomonas species is discussed. Received: 17 June 1998 / Accepted: 12 October 1998     

Identification and expression of a sialyltransferase responsible for the synthesis of disialylgalactosylgloboside in normal and malignant kidney cells: downregulation of ST6GalNAc VI in renal cancers

Although disialyl glycosphingolipids such as GD3 and GD2 have been considered to be associated with malignant tumours, whether branched-type disialyl glycosphingolipids show such an association is not well understood. We investigated the sialyltransferases responsible for the biosynthesis of DSGG (disialylgalactosylgloboside) from MSGG (monosialylgalactosylgloboside). Among six GalNAc:alpha2,6-sialyltransferases cloned to date, we focused on ST6GalNAc III, V and VI, which utilize sialylglycolipids as substrates. In vitro enzyme analyses revealed that ST6GalNAc III and VI generated DSGG from MSGG with V(max)/K(m) values of 1.91 and 4.16 respectively. Transfection of the cDNA expression vectors for these enzymes resulted in DSGG expression in a renal cancer cell line. Although both ST6GalNAc III and VI genes were expressed in normal kidney cells, the expression profiles of ST6GalNAc VI among 20 renal cancer cell lines correlated clearly with those of DSGG, suggesting that the sialyltransferase involved in the synthesis of DSGG in the kidney is ST6GalNAc-VI. ST6GalNAc-VI and DSGG were found in proximal tubule epithelial cells in normal kidney tissues, while they were downregulated in renal cancer cell lines and cancer tissues. All these findings indicated that DSGG was suppressed during the malignant transformation of the proximal tubules as a maturation arrest of glycosylation.     

Influence of oxygen on the growth of Saccharomyces cerevisiae in continuous culture

Baker's yeast, Saccharomyces cerevisiae, was investigated for the combined influence of dissolved oxygen and glucose concentration in continuous culture. A reactor was operated at a range of dilution rates (0.1, 0.2, 0.25, 0.27, and 3.0 h(-1)), above and below the critical value that separates the oxidative and fermentation regions. For each dilution rate (D), steady states were established at each of five to ten different dissolved oxygen concentrations (DO) in the range of 0.01-5 mg/L. The use of on-line mass spectrometry facilitated the measurement of gaseous and dissolved O(2), CO(2), and ethanol. Intracellular carbohydrate, protein, RNA, DNA, lipid, and cytochrome concentrations were measured. Cell size measurements were reduced to specific surface areas. Cytochrome content showed up to 100% variation during a 20-day period of adaptation at D = 0.2 h(-1) to low DO. Eventually, the culture behaved the same at DO = 0.05 mg/L as it did initially at 3 mg/L. At D = 0.2, 0.25, and 0.27 h(-1), the transition between oxidation and fermentation was characterized by a critical DO which decreased with decreasing D. The X-D curves were shifted such that the critical D value was reduced with decreasing DO. Specific oxygen update rates varied with DO according to the saturation kinetics. Specific cell surface areas increased with decreasing DO. Cytochrome content generally decreased with decreasing DO, and Q(O(2) ) could be linearly related to the total cytochrome content, which exhibited a maximum at D = 0.27 h(-1).     

Factors influencing lifespan dependency on agricultural crops by brown bears

Landscape and Ecological Engineering - Investigating factors underlying human-wildlife conflicts in agricultural landscapes is important for both preventing crop damage and wildlife conservation....     

Evaluation of tri-combinant vaccine for feline herpesvirus, calicivirus and panleukopenia virus infections in Japanese native cats

Tri-combinant vaccine consisting of attenuated feline herpesvirus (FHV) and feline calicivirus (FCV) and inactivated feline panleukopenia virus (FPLV), were evaluated for safety and efficacy, using Japanese native cats and the viral strains isolated in Japan. Thirty-eight 9- to 12-week-old kittens were inoculated intramuscularly and subcutaneously with the vaccine. Consequently, no adverse reaction was found, and protective efficacy was confirmed by challenge tests with the virulent strains of each virus. Serum-neutralizing antibodies against FCV and FPLV were maintained for at least one year after vaccination, whereas antibody against FHV disappeared in two cases at 24 weeks after vaccination. Application of this vaccine seemed effective for control of feline viral disease in cats for experimental use.     

Nitric Oxide-Induced [3H]GABA Release from Cerebral Cortical Neurons Is Mediated by Peroxynitrite

Abstract: The functional significance of peroxynitrite in the release of [³H]GABA induced by nitric oxide (NO) liberated from NO generators was investigated using cerebral cortical neurons in primary culture. NO generators such as sodium nitroprusside (SNP) and S -nitroso- N -acetylpenicillamine (SNAP) increased [³H]GABA release in a dose-dependent manner. These increases in [³H]GABA release were significantly inhibited by hemoglobin, indicating that those NO generators evoke the release of [³H]GABA by the formation of NO. Two types of superoxide scavengers, Cu²⁺/Zn²⁺ superoxide dismutase and ceruloplasmin, significantly reduced the increase in [³H]GABA release induced by both SNP and SNAP, which assumes that NO requires superoxide to induce [³H]GABA release from the neurons. In addition, synthesized peroxynitrite induced a dose-dependent increase in [³H]GABA release from the neurons. These results indicate that NO-induced [³H]GABA release is mediated by peroxynitrite formed by the reaction of NO with superoxide.     

Rapid startup and high rate nitrogen removal from anaerobic sludge digester liquor using a SNAP process

In this study, a single-stage autotrophic nitrogen removal reactor, packed with a novel acrylic fiber biomass carrier material (Biofix), was applied for nitrogen removal from sludge digester liquor. For rapid start-up, conventional activated sludge was added to the reactor soon after the attachment of anammox biomass on the Biofix carriers, which allowed conventional activated sludge to form a protective layer of biofilm around the anammox biomass. The Nitrogen removal efficiency reached 75% within 1 week at a nitrogen loading rate of 0.46 kg-N/m³/day for synthetic wastewater treatment. By the end of the synthetic wastewater treatment period, the maximum nitrogen removal rate had increased to 0.92 kg-N/m³/day at a nitrogen loading rate of 1.0 kg-N/m³/day. High nitrogen removal rate was also achieved during the actual raw digester liquor treatment with the highest nitrogen removal rate being 0.83 kg-N/m³/day at a nitrogen loading rate of 0.93 kg-N/m³/day. The thick biofilm on Biofix carriers allowed anammox bacteria to survive under high DO concentration of 5–6 mg/l resulting in stable and high nitrogen removal performance. FISH and CLSM analysis demonstrated that anammox bacteria coexisted and surrounded by ammonium oxidizing bacteria.     

Unique Induction of CA1 LTP Components After Intake of Theanine, an Amino Acid in Tea Leaves and its Effect on Stress Response

Theanine, γ-glutamylethylamide, is one of the major amino acid components in green tea. This study was undertaken to evaluate the effect of theanine intake on long-term potentiation (LTP) induction at hippocampal CA1 synapses and exposure to acute stress. Young rats were fed water containing 0.3% theanine after birth. Key findings: Serum corticosterone level was markedly decreased by theanine intake. Because this decrease can modify synaptic plasticity, the effect of theanine intake was examined focused on CA1 LTP induction. CA1 LTP induced by a 100-Hz tetanus for 1 s was almost the same extent in hippocampal slices from theanine-administered rats, whereas that induced by a 200-Hz tetanus for 1 s was significantly attenuated. 2-Amino-5-phosphonovalerate (APV), an N-methyl-d-aspartate (NMDA) receptor antagonist, significantly attenuated CA1 LTP induced by a 200-Hz tetanus in the control rats, but not in theanine-administered rats. Interestingly, APV completely blocked CA1 LTP induced by a 100-Hz tetanus in the control rats, while scarcely blocking it in theanine-administered rats. These results indicate that theanine intake reduces NMDA receptor-dependent CA1 LTP, while increasing NMDA receptor-independent CA1 LTP. Furthermore, neither 100-Hz tetanus-induced LTP nor 200-Hz tetanus-induced LTP was attenuated in theanine-administered rats after exposure to tail suspension stress, suggesting that the lack of NMDA receptor-dependent CA1 LTP by theanine intake is involved in ameliorating the attenuation of CA1 LTP after tail suspension. This study is the first to indicate that theanine intake modifies the mechanism of CA1 LTP induction.