Ammonia assimilation in Klebsiella pneumoniae F-5-2 that can utilize ammonium and nitrate ions simultaneously: purification and characterization of glutamate dehydrogenase and glutamine synthetase

The two ammonia-assimilating enzymes glutamate dehydrogenase (GDH; EC 1.4.1.4) and glutamine synthetase (GS; EC 6.3.1.2) were synthesized steadily during the cell growth of Klebsiella pneumoniae F-5-2 that can utilize NH4+ and NO3- simultaneously under aerobic conditions. The enzymes were purified to homogeneity from cell extracts and characterized. The molecular mass of the purified GDH was 300 kDa with six identical 52-kDa subunits. GDH showed its maximal activity (aminating) at pH 8.0 and was stable between pHs 5.5 and 11.5. The enzyme was NADP-specific and strongly inhibited by Ag+. It catalyzed the amination of 2-ketovalerate, 2-ketoadipate, and 2-ketobutyrate, in addition to 2-ketoglutarate. The purified GS has a molecular mass of 470 kDa with eight identical 60-kDa subunits. GS showed its maximal activity at pH 8.0 and was stable between pHs 6.0 and 7.0. The enzyme was strongly inhibited by Fe3+, Hg2+, and Cu2+.     

Stability tests and stabilization for piecewise linear systems based on poles and zeros of subsystems

This paper provides several stability tests for piecewise linear systems and proposes a method of stabilization for bimodal systems. In particular, we derive an explicit and exact stability test for planar systems, which is given in terms of coefficients of transfer functions of subsystems. Restricting attention to the bimodal and planar case, we show simple stability tests. In addition, we drive a necessary stability condition and a sufficient stability condition for higher-order and bimodal systems. They are given in terms of the eigenvalue loci and the observability of subsystems. All the stability tests provided in this paper are computationally tractable, and our results are applied to the stabilizability problem. We confirm the exactness and effectiveness of our approach by illustrative examples.     

Spontaneous hydrolytic degradability of copolyesters having tetrahydropyran rings in their backbones

Homo- and copolyesters, containing 2,5-linked tetrahydropyran rings in their backbones, were prepared by cationic ring-opening polymerization of bicyclic lactones, 2,6-dioxabicyclo [2.2.2] octan-3-one ( 1 ), and its 4-methoxy-carbonyl and 4-methoxy-carbonyl and 4-benzyloxycarbonyl derivatives ( 3 and 4 ). Copolyesters, having pendant carboxyl groups, were derived from the copolymers containing 4 as one of the comonomers by the catalytic hydrogenolysis of the pendant benzyloxy–carbonyl groups. Copolyesters, containing both 2,5-linked and 2,6-linked tetrahydropyran rings in the main chains, were obtained by the copolymerization of 1 with 6,8-dioxabicyclo [3.2.1] octan-7-one ( 13 ), which is a structural isomer of 1 . Spontaneous hydrolytic degradability of these polyesters was investigated in a phosphate buffer solution (pH 7.5) at 27°C. The degradability of the polyesters markedly depended on the molecular structure: hydrophobic pendant groups retarded the hydrolytic degradation, whereas hydrophilic pendant groups accelerated it. In particular, a small amount of pendant carboxyl groups significantly enhanced the hydrolytic degradability of the copolyesters from 1 . Polyesters, containing 2,6-linked tetrahydropyran rings in their backbones, underwent hydrolytic degradation more readily than those entirely consisting of 2,5-linked tetrahydropyran rings. © 1994 John Wiley & Sons, Inc.     

Far-infrared laser oscillation due to cyclotron emission in p-type germanium

Far-infrared laser oscillation due to cyclotron emission in the light hole band of p-type germanium was observed under crossed electric and magnetic fields. The wavelength is inversely proportional to magnetic fields with a cyclotron mass ofm _c =0.048m _c . Numerical calculations based on Luttinger Hamiltonian show that mixing of wavefunctions between the light and heavy hole bands causes population inversion betweenn=0 andn=1 light-hole Landau levels. It is also shown that non-equidistant energy spacing of light-hole Landau levels is essential to yield net amplification.     

Requirement for activation of the serine-threonine kinase Akt (protein kinase B) in insulin stimulation of protein synthesis but not of glucose transport

A wide variety of biological activities including the major metabolic actions of insulin is regulated by phosphatidylinositol (PI) 3-kinase. However, the downstream effectors of the various signaling pathways that emanate from PI 3-kinase remain unclear. Akt (protein kinase B), a serine-threonine kinase with a pleckstrin homology domain, is thought to be one such downstream effector. A mutant Akt (Akt-AA) in which the phosphorylation sites (Thr308 and Ser473) targeted by growth factors are replaced by alanine has now been shown to lack protein kinase activity and, when overexpressed in CHO cells or 3T3-L1 adipocytes with the use of an adenovirus vector, to inhibit insulin-induced activation of endogenous Akt. Akt-AA thus acts in a dominant negative manner in intact cells. Insulin-stimulated protein synthesis, which is sensitive to wortmannin, a pharmacological inhibitor of PI 3-kinase, was abolished by overexpression of Akt-AA without an effect on amino acid transport into the cells, suggesting that Akt is required for insulin-stimulated protein synthesis. Insulin activation of p70 S6 kinase was inhibited by approximately 75% in CHO cells and approximately 30% in 3T3-L1 adipocytes, whereas insulin-induced activation of endogenous Akt was inhibited by 80 to 95%, by expression of Akt-AA. Thus, Akt activity appears to be required, at least in part, for insulin stimulation of p70 S6 kinase. However, insulin-stimulated glucose uptake in both CHO cells and 3T3-L1 adipocytes was not affected by overexpression of Akt-AA, suggesting that Akt is not required for this effect of insulin. These data indicate that Akt acts as a downstream effector in some, but not all, of the signaling pathways downstream of PI 3-kinase.     

A robust increase in expression of arc gene, an effector immediate early gene, in the rat brain after acute and chronic methamphetamine administration

The effect of acute and chronic administration of methamphetamine (METH) on the levels of activity-regulated cytoskeleton-associated protein (arc), an effector-immediate early gene, mRNA has been investigated in rat brain using in situ hybridization. Levels of arc mRNAs in the brain regions examined increased significantly from 0.5-1 h after an acute METH (4 mg/kg) administration compared with basal levels. The increase in arc mRNA continued by 3 h, and then subsided to basal levels by 6 h. The degree of increase in arc mRNA and the peak time after METH administration varied according to brain area. Arc mRNA in cerebral cortices showed robust increase 1 h after METH administration. In the striatum and hippocampus, it showed earlier and later increase, respectively, and its degree of both was less than in the cortices. Microscopic examination revealed that the METH-induced arc mRNAs in the parietal cortex were enriched in layers IV and VI, and those in the striatum existed mainly in the medium-sized neuron. Pretreatment with either 0.5 mg/kg SCH23390 or 0.25 mg/kg MK-801 almost completely blocked the enhanced striatal arc mRNA levels induced by acute METH administration, whereas such pretreatments only partially reduced the effect of METH in the cerebral cortical regions. In the chronic treatment experiment, the arc mRNA levels of the group that received chronic treatment with METH followed by a METH challenge showed an increase like seen after acute METH administration. Since previous studies proposed that arc is one of cytoskeleton-associated proteins and is selectively localized in neural dendrites, the results of the present study suggested that arc may play an important role in the synaptic plasticity underlying METH-induced adaptational changes including behavioral sensitization.     

Efficacy of dose-intensified MEC (methotrexate, epirubicin and cisplatin) chemotherapy for advanced urothelial carcinoma: a prospective randomized trial comparing MEC and M-VAC (methotrexate, vinblastine, doxorubicin and cisplatin). Japanese Urothelial Cancer Research Group

BACKGROUND: To evaluate the antitumor activity in patients with T3b, T4 or metastatic urothelial carcinoma treated with MEC or M-VAC chemotherapy, by performing a multi-center randomized prospective study. METHODS: From 1991 to 1995, 89 patients with T3b, T4 or metastatic urothelial carcinoma were randomly allocated to a methotrexate, epirubicin and cisplatin chemotherapy group (arm 1: S-MEC therapy; n = 29), a dose-intensified MEC therapy combined with G-CSF group (arm 2: I-MEC therapy; n = 30) or a methotrexate, vinblastine, doxorubicin and cisplatin chemotherapy (arm 3: M-VAC therapy; n = 30). At the registration center, the patients were stratified into previously untreated patients and patients with recurrence after radical operation and then randomly allocated to the treatment groups. In each arm, two or more courses of chemotherapy (4-week cycles) were performed. RESULTS: Of the 88 eligible patients, four treated with S-MEC therapy and two treated with I-MEC therapy showed CR. The response rates (CR + PR) were 52% (15/29) with S-MEC therapy, 76% (22/29) with I-MEC therapy and 47% (14/30) with M-VAC therapy. The response rate with I-MEC therapy was significantly higher than that with M-VAC therapy (P = 0.02). Although the incidence of leukopenia was low with I-MEC therapy, the incidence of thrombocytopenia was high with this therapy. CONCLUSION: MEC therapy used in this study is promising in terms of the antitumor effects.     

Recombinant modified vaccinia virus Ankara-simian immunodeficiency virus gag pol elicits cytotoxic T lymphocytes in rhesus monkeys detected by a major histocompatibility complex class I/peptide tetramer

The utility of modified vaccinia virus Ankara (MVA) as a vector for eliciting AIDS virus-specific cytotoxic T lymphocytes (CTL) was explored in the simian immunodeficiency virus (SIV)/rhesus monkey model. After two intramuscular immunizations with recombinant MVA-SIVSM gag pol, the monkeys developed a Gag epitope-specific CTL response readily detected in peripheral blood lymphocytes by using a functional killing assay. Moreover, those immunizations also elicited a population of CD8+ T lymphocytes in the peripheral blood that bound a specific major histocompatibility complex class I/peptide tetramer. These Gag epitope-specific CD8+ T lymphocytes also were demonstrated by using both functional and tetramer-binding assays in lymph nodes of the immunized monkeys. These observations suggest that MVA may prove a useful vector for an HIV-1 vaccine. They also suggest that tetramer staining may be a useful technology for monitoring CTL generation in vaccine trials in nonhuman primates and in humans.     

Modification of parallel activity elicited by propagating bursts in developing networks of rat cortical neurones

The fine specificity of the cellular immune response to Candida albicans (i.e., recognition of different antigenic components) between normal controls and human immunodeficiency virus-infected patients in various stages of disease was compared. C. albicans-specific T cells, enriched by antigen stimulation and interleukin-2 expansion, were challenged with antigenic fractions of different molecular weight obtained by SDS-gel fractionation of C. albicans extracts in the presence of autologous mononuclear cells as antigen-presenting cells. Proliferative responses showed similar patterns of reactivity between controls and category A and B seropositive subjects. Category C patients with concurrent C. albicans infections did not give rise to C. albicans-specific T cell lines, confirming the T cell defect. Patients without clinically evident C. albicans infection had a low but broad reactivity pattern of C. albicans-specific T cells. These results suggest that depletion of C. albicans-specific T cells, independent of their fine specificity, occurs along with disease progression.     

Measurement and correlation of mutual diffusion coefficients for styrene-butadiene rubber(SBR)-n-nonane systems

The mutual diffusion coefficients for styrene-butadiene rubber(SBR)-n-nonane systems were measured using a sorption apparatus with a quartz spring at 100 and 130°C for mass fractions of n-nonane ranging from 0 to 0.16. The mutual diffusion coefficients were correlated with good agreement by the free-volume theory.