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At least 649 records · Page 36Linked to original sources
PubMed1993

Glutamate synthase is plastid-encoded in a red alga: implications for the evolution of glutamate synthases.

An actively transcribed gene (glsF) encoding for ferredoxin-dependent glutamate synthase (Fd-GOGAT) was found on the plastid genome of the multicellular red alga Antithamnion sp. Fd-GOGAT is not plastid-encoded in chlorophytic plants, demonstrating that red algal plastid genomes encode for additional functions when compared to those known from green chloroplasts. Moreover, our results suggest that the plant Fd-GOGAT has an endosymbiotic origin. The same may not be true for NADPH-dependent GOGAT. In Antithamnion glsF is flanked upstream by cpcBA and downstream by psaC and is transcribed monocistronically. Implications of these results for the evolution of GOGAT enzymes and the plastid genome are discussed.

Amino Acid Sequence↗
PubMed2005

Self-association of glutamic acid-rich fusion peptide analogs of influenza hemagglutinin in the membrane-mimic environments: effects of positional difference of glutamic acids on side chain ionization constant and intra- and inter-peptide interactions deduced from NMR and gel electrophoresis measurements.

Two glutamic acid-rich fusion peptide analogs of influenza hemagglutinin were synthesized to study the organization of the charged peptides in the membranous media. Fluorescence and gel electrophoresis experiments suggested a loose association between the monomers in the vesicles. A model was built which showed that a positional difference of 3, 7 and 4, 8 results in the exposure of Glu3 and Glu7 side chains to the apolar lipidic core. Supportive results include: first, pK(a) values of two pH units higher than reference value in aqueous medium for Glu3 and Glu7 CgammaH, whereas the deviation of pK(a) from the reference value for Glu4 and Glu8 CgammaH is substantially smaller; second, Hill coefficients of titration shift of these protons indicate anti-cooperativity for Glu3 and Glu7 side chain protons but less so for Glu4 and Glu8, implying a strong electrostatic interaction between Glu3 and Glu7 possibly resulting from their localization in an apolar environment; third, positive and larger titration shift for NH of Glu3 is observed compared to that of Glu4, suggesting stronger hydrogen bond between the NH and the carboxylic group of Glu3 than that of Glu4, consistent with higher degree of exposure to hydrophobic medium for the side chain of Glu3.

Caprylates↗
PubMed2005

Effects of glutamate transporter and receptor ligands on neuronal glutamate uptake.

The excitatory amino acids (EAAs) transporters regulate the balance between physiological and pathological signaling over stimulation of the glutamatergic system pathway. The effect of transportable substrates and glutamate (Glu) receptor agonists on Glu uptake in neuronal cells was assessed at different conditions. Cells pre-incubated with Glu, L- or D-aspartate (Asp) and washed presented an inhibition on [(3)H]-Glu uptake and this effect was not mimicked by Glu receptors agonists. The effects of L- and D-Asp were not altered by the presence of N-methyl-d-aspartate (NMDA) receptor antagonists. Thus, the reduction on Glu uptake induced by EAAs is probably linked to the transporter activity. In contrast, the presence of NMDA or (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (SR-ACPD) during the pre-incubation and the [(3)H]-Glu uptake assay period increased Glu uptake, whilst kainic acid (KA) had no effect. The NMDA effect was not altered by its antagonists (+/-)-2-amino-5-phosphonopentanoic acid (AP-5) or dizocilpine (MK-801). The SR-ACPD effect was due to the activation of metabotropic Glu receptor, since it was abolished by its antagonist, L(+/-)-2-amino-3-phosphonopropionic acid (L-AP3). Thus, the current studies suggest that the neuronal EAAs transporter is regulated in different manner by transportable substrates and Glu receptor agonists. The possible involvement of this modulation after certain neurotoxicity insults is discussed.

Amino Acid Transport System X-AG↗
PubMed2004

Synaptically stimulated induction of group I metabotropic glutamate receptor-dependent long-term depression and depotentiation is inhibited by prior activation of metabotropic glutamate receptors and protein kinase C.

We have investigated metaplasticity of the group I metabotropic glutamate receptor (mGluR)-dependent long-term depression (LTD) and depotentiation (DP) induced by physiological synaptic stimulation in the medial perforant path of the dentate gyrus in vitro. Group I mGluR-LTD/DP was inhibited by prior preconditioning brief high frequency stimulation (HFS) if the preconditioning HFS induced long-term potentiation (LTP) or if the induction of LTP was inhibited by an NMDA receptor antagonist. The inhibitory effect of the preconditioning HFS on LTD/DP was dependent upon activation of mGluRs, as it was blocked by the presence of the mGluR antagonist (S)-alpha-methyl-4-carboxyphenylglycine during the preconditioning stimulation. The inhibitory effect of the preconditioning HFS involved stimulation of PKC, as the presence of the PKC inhibitor bisindolylmaleimide (BIS) during the preconditioning stimulation prevented the inhibitory effect of such preconditioning stimulation. Activation of PKC was also necessary for the induction of mGluR-LTD itself, as the PKC inhibitor BIS prevented the induction of the mGluR-LTD. We suggest that the physiological stimulation of mGluRs by the preconditioning stimulation produces a PKC-dependent inactivation of subsequent group I mGluR functioning and thereby an inhibition of induction of group I mGluR-dependent LTD/DP induction.

Animals↗
PubMed1998

The effects of monosodium glutamate in adults with asthma who perceive themselves to be monosodium glutamate-intolerant.

BACKGROUND: Many previous clinical studies of food-induced asthma suffer from inadequate baseline or control data. A statistically valid, randomized, double-blind, placebo-controlled, monosodium glutamate (MSG)-challenge protocol was developed for identifying early and late asthmatic reactions in an individual. OBJECTIVE: We sought to determine whether MSG would induce bronchoconstriction in a group of adults with asthma who perceived that they were MSG sensitive. METHODS: Twelve subjects (seven women, mean age 35.3 years) with clinically documented asthma and a perception of MSG-induced asthma were recruited. FEV1 and peak expiratory flow data were obtained for 3 whole control days, as well as time-matched data for 3 separate challenge days (1 gm MSG, 5 gm MSG, and 5 gm lactose [placebo]). Opaque capsule challenges were given as a single dose in the morning after an overnight fast. Subjects complied with an elimination diet throughout the study. Nonspecific bronchial hyperresponsiveness was measured at baseline, after the control days, and at the conclusion of the challenges. Venous blood samples were taken at baseline and on each challenge day to determine soluble inflammatory marker (eosinophil cationic protein and tryptase) activity. RESULTS: No immediate or definite late asthmatic reactions occurred. One subject's FEV1 declined more than 15% on MSG challenge, but 95% confidence limits for the control-day spirometry showed that this decline was within her daily variation, hence the challenge was deemed to be negative. No significant changes in bronchial hyperresponsiveness or soluble inflammatory markers were found. CONCLUSIONS: MSG-induced asthma was not demonstrated in this study. This study highlighted the importance of adequate baseline and control data and indicated that such a rigorous protocol for individual assessment is feasible.

Adult↗
PubMed1997

Glutamate toxicity in neuron-enriched and neuron-astrocyte co-cultures: effect of the glutamate uptake inhibitor L-trans-pyrrolidine-2,4-dicarboxylate.

Exposure of neuron-enriched cultures of the embryonic rat cerebral cortex to glutamate (Glu) resulted in a concentration-dependent (EC50 = 50 microM) increase in the release of lactate dehydrogenase (LDH) into the bathing medium. Glu-induced neurotoxicity appeared to be mediated predominantly by the activation of N-methyl-D-aspartate receptors because its effects were almost completely reversed by 1 microM (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine hydrogen maleate (MK-801). The potency of Glu was increased by ca eight-fold (EC50 = 6 microM) when neuronal cultures were incubated with Glu in the presence of the uptake inhibitor L-trans-pyrrolidine-2, 4-dicarboxylate (L-trans-PDC). Increased LDH release was also observed when neuron-enriched cultures were exposed to an ineffective concentration of Glu in the presence of increasing concentrations of L-trans-PDC, moreover, the uptake inhibitor itself became neurotoxic at concentrations above 100 microM. Glu also stimulated the release of LDH from neuron-astrocyte co-cultures in a concentration-dependent manner, however, it was found to be less potent (EC50 = 200 microM) in these cultures than it was in the neuron-enriched cultures. Incubation of the cocultures with Glu in the presence of the uptake inhibitor increased the potency of Glu (EC50 = 100 microM) but to a much lesser extent than that found in neuron-enriched cultures. Cultured astrocytes were found to be resistant to injury by Glu even in the presence of the uptake inhibitor.

Animals↗
PubMed1983

Glutamic acid decarboxylase in cerebrospinal fluid in infancy and childhood Part II. Glutamic acid decarboxylase activity in cerebrospinal fluid of children with neurological diseases.

Glutamic acid decarboxylase (GAD) activity in cerebrospinal fluid (CSF) was determined in 53 patients with neurological diseases as follows: Epilepsy (n:17), febrile convulsions (n:3), meningoencephalitis (n:17), encephalopathies (n:10), CNS leukemia (n:3), congenital hydrocephalus (n:2) and pseudoileus neonatorum (n:1). Compared with the mean normal value (5.2 +/- 2.5 pmol CO2 formed/hr/ml) reported in Part I, a significant increase of GAD activity in CSF was demonstrated in patients with uncontrolled epileptic seizures (11.4 +/- 3.9 pmol CO2 formed/hr/ml), febrile convulsions (13.5 +/- 8.7), viral meningitis with or without encephalitis (20.3 +/- 13.6), encephalopathies (30.0 +/- 25.9), CNS leukemia (11.1 +/- 5.0), congenital hydrocephalus (20.5 +/- 7.3) and pseudoileus neonatorum (28.6). Markedly high GAD activity was found in patients with CNS leukemia several days after intrathecal injection of methotrexate (39.8 +/- 18.0). On the other hand, significantly low GAD activity was shown in patients with bacterial meningitis or brain abscess (1.3 +/- 1.2). This suggests that some bacterial factors may be inhibitory toward GAD activity in CSF. High GAD activity in CSF may be useful as an indicator of aseptic brain dysfunction, although it was not always correlated with the severity of symptoms.

Adolescent↗
PubMed1994

Site-directed mutagenesis of the catalytic base glutamic acid 400 in glucoamylase from Aspergillus niger and of tyrosine 48 and glutamine 401, both hydrogen-bonded to the gamma-carboxylate group of glutamic acid 400.

Replacement of the catalytic base Glu400 by glutamine in glucoamylase from Aspergillus niger affects both substrates ground-state binding and transition-state stabilization. Compared to those of the wild-type enzyme, Km values for maltose and maltoheptaose are 12- and 3-fold higher for the Glu400-->Gln mutant, with kcat values 35- and 60-fold lower, respectively, for the same substrates. This unusually high residual activity for a glycosylase mutant at a putative catalytic group is tentatively explained by a reorganization of the hydrogen bond network, using the crystal structure of the related Aspergillus awamori var. X100 glucoamylase in complex with 1-deoxynojirimycin [Harris, E. M. S., Aleshin, A. E., Firsov, L. M., & Honzatko, R. B. (1993) Biochemistry 32, 1618-1626]. Supposedly Gln400 in the mutant hydrogen bonds to the invariant Tyr48, as does Glu400 in the wild-type enzyme. For Tyr48-->Trp A. niger glucoamylase kcat is reduced 80-100-fold, while Km is increased only 2-3-fold. Gln401 also hydrogen bonds to Glu400, but its mutation to glutamic acid has only a minor effect on activity. The Tyr48-->Trp and Glu400-->Gln glucoamylases share particular features in displaying unusually high activity below pH 4.0-which reflects lack of the wild-type catalytic base function- and unusually low binding affinity at subsite 2. Both mutants have lost 13-16 kJ mol-1 in transition-state stabilization energy.(ABSTRACT TRUNCATED AT 250 WORDS)

Aspergillus niger↗
PubMed1999

Role of glutamate-59 hydrogen bonded to N(3)H of the flavin mononucleotide cofactor in the modulation of the redox potentials of the Clostridium beijerinckii flavodoxin. Glutamate-59 is not responsible for the pH dependency but contributes to the stabilization of the flavin semiquinone.

The midpoint potentials for both redox couples of the noncovalently bound flavin mononucleotide (FMN) cofactor in the flavodoxin are known to be pH dependent. While the pH dependency for the oxidized-semiquinone (ox/sq) couple is consistent with the formation of the blue neutral form of the flavin semiquinone, that of the semiquinone-hydroquinone (sq/hq) couple is more enigmatic. The apparent pK(a) of 6.7 for this couple in the flavodoxin from Clostridium beijerinckii has been attributed to the ionization of the FMN(HQ); however, nuclear magnetic resonance data strongly suggest the FMN(HQ) remains anionic over the entire pH range testable. As an alternative explanation, a specific glutamate residue (Glu59 in this flavodoxin), which is hydrogen-bonded to N(3)H of the FMN, has been postulated to be the primary redox-linked proton acceptor responsible for the pH effect in some flavodoxins. This model was directly tested in this study by permanently neutralizing Glu59 by its replacement with glutamine. This conservative substitution resulted in an increase of 86 mV (at pH 7) in midpoint potential of the sq/hq couple; however, the pH dependency of this couple was not altered. Thus, the redox-linked protonation of Glu59 clearly cannot be responsible for this effect as proposed. The pH dependency of the ox/sq couple was also similar to wild type, but the midpoint potential has decreased by 65 mV (pH 7). The K(d) values for the oxidized, semiquinone, and hydroquinone complexes increased by 43-, 590-, and 20-fold, respectively, relative to the wild type. Thus, the Glu59 to glutamine substitution substantially effects the stability of the semiquinone but, on a relative basis, slightly favors the formation of the hydroquinone. On the basis of (1)H-(15)N HSQC nuclear magnetic resonance spectroscopic studies, the increased temperature coefficients for the protons on N(3) and N(5) of the reduced FMN in E59Q suggest that the hydrogen-bonding interactions at these positions are significantly weakened in this mutant. The increase for N(5)H correlates with the reduced stability of the FMN(SQ) and the more negative midpoint potential for the ox/sq couple. On the basis of the X-ray structure, an "anchoring" role is proposed for the side chain carboxylate of Glu59 that stabilizes the structure of the 50's loop in such a way so as to promote the crucial hydrogen-bonding interaction that stabilizes the flavin semiquinone, contributing to the low potential of this flavodoxin.

Circular Dichroism↗
PubMed2001

Phosphinic acid pseudopeptides analogous to glutamyl-gamma-glutamate: synthesis and coupling to pteroyl azides leads to potent inhibitors of folylpoly-gamma-glutamate synthetase.

Several routes to a complex phosphinate phosphapeptide analogous to the gamma-glutamyl peptide Glu-gamma-Glu have been investigated. Formation of gamma-phosphono glutamate derivatives via addition of a phosphorus-based radical to protected vinylglycine was found to be of limited value because of the elevated temperatures required. Alkylation and conjugate addition reactions of trivalent phosphorus (P(III)) species were investigated. In situ generation of bis-trimethylsilyl esters of phosphinous acids proved to be an effective route to phosphinates of modest structural complexity. However, this chemistry could not be extended to the incorporation of an amino acid moiety at the N-terminal side of the desired phosphinate. A successful synthesis of the target phosphinate phosphapeptide was effected using P(III) chemistry and dehydrohalogenation to yield an alpha,beta-unsaturated phosphinic acid ester, following which conjugate addition of diethylacetamido malonate and acid-mediated hydrolysis afforded the desired phosphinate phosphapeptide. Coupling of the unprotected phosphinate phosphapeptide with two acyl azides derived from folic acid and methotrexate led to the corresponding pteroylphosphapeptides of interest as possible mimics of tetrahedral intermediates in the reaction catalyzed by folylpolyglutamate synthetase.

Azides↗
PubMed2000

NADP+-dependent glutamate dehydrogenase in the Antarctic psychrotolerant bacterium Psychrobacter sp. TAD1. Characterization, protein and DNA sequence, and relationship to other glutamate dehydrogenases.

The Antarctic psychrotolerant bacterium Psychrobacter sp. TAD1 contains two distinct glutamate dehydrogenases (GDH), each specific for either NADP+ or NAD+. This feature is quite unusual in bacteria, which generally have a single GDH. NADP+-dependent GDH has been purified to homogeneity and the gene encoding GDH has been cloned and expressed. The enzyme has a hexameric structure. The amino acid sequence determined by peptide and gene analyses comprises 447 residues, yielding a protein with a molecular mass of 49 285 Da. The sequence shows homology with hexameric GDHs, with identity levels of 52% and 49% with Escherichia coli and Clostridium symbiosum GDH, respectively. The coenzyme-binding fingerprint motif GXGXXG/A (common to all GDHs) has Ser at the last position in this enzyme. The overall hydrophilic character is increased and a five-residue insertion in a loop between two alpha-helices may contribute to the increase in protein flexibility. Psychrobacter sp. TAD1 GDH apparent temperature optimum is shifted towards low temperatures, whereas irreversible heat inactivation occurs at temperatures similar to those of E. coli GDH. The catalytic efficiency in the temperature range 10-30 degrees C is similar or lower than that of E. coli GDH. Unlike E. coli GDH the enzyme exhibits marked positive cooperativity towards 2-oxoglutarate and NADPH. This feature is generally absent in prokaryotic GDHs. These observations suggest a regulatory role for this GDH, the most crucial feature being the structural/functional properties required for fine regulation of activity, rather than the high catalytic efficiency and thermolability encountered in several cold-active enzymes.

Amino Acid Motifs↗
PubMed2001

Glutamic acid decarboxylase and glutamate receptor changes during tolerance and dependence to benzodiazepines.

Protracted administration of diazepam elicits tolerance, whereas discontinuation of treatment results in signs of dependence. Tolerance to the anticonvulsant action of diazepam is present in an early phase (6, 24, and 36 h) but disappears in a late phase (72-96 h) of withdrawal. In contrast, signs of dependence such as decrease in open-arm entries on an elevated plus-maze and increased susceptibility to pentylenetetrazol-induced seizures were apparent 96 h (but not 12, 24, or 48 h) after diazepam withdrawal. During the first 72 h of withdrawal, tolerance is associated with changes in the expression of GABA(A) (gamma-aminobutyric acid type A) receptor subunits (decrease in gamma(2) and alpha(1); increase in alpha(5)) and with an increase of mRNA expression of the most abundant form of glutamic acid decarboxylase (GAD), GAD(67). In contrast, dl-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor GluR1 subunit mRNA and cognate protein, which are normal during the early phase of diazepam withdrawal, increase by approximately 30% in cortex and hippocampus in association with the appearance of signs of dependence 96 h after diazepam withdrawal. Immunohistochemical studies of GluR1 subunit expression with gold-immunolabeling technique reveal that the increase of GluR1 subunit protein is localized to layer V pyramidal neurons and their apical dendrites in the cortex, and to pyramidal neurons and in their dendritic fields in hippocampus. The results suggest an involvement of GABA-mediated processes in the development and maintenance of tolerance to diazepam, whereas excitatory amino acid-related processes (presumably via AMPA receptors) may be involved in the expression of signs of dependence after withdrawal.

Animals↗
PubMed1995

Targeting of the 67-kDa isoform of glutamic acid decarboxylase to intracellular organelles is mediated by its interaction with the NH2-terminal region of the 65-kDa isoform of glutamic acid decarboxylase.

The two isoforms of glutamic acid decarboxylase (GAD), GAD67 and GAD65, synthesize the neurotransmitter gamma-aminobutyric acid in neurons and pancreatic beta-cells. Previous studies suggest that GAD67 is a soluble cytosolic protein, whereas GAD65 is membrane-associated. Here, we study the intracellular distribution of GAD67 in neurons, pancreatic beta-cells, and fibroblasts transfected either with GAD65 and GAD67 together or with GAD67 alone. Neuronal GAD67 is partially recovered with GAD65 in membrane-containing pellet fractions and Triton X-114 detergent phases. The two proteins co-immunoprecipitate from extracts of brain and GAD65-GAD67 co-transfected fibroblasts, but not when extracts of GAD65 and GAD67 transfected fibroblasts were mixed and used as a starting material for immunoprecipitation. GAD67 is concentrated in the Golgi complex region in GAD65-GAD67 co-transfected fibroblasts, but not in fibroblasts transfected with GAD67 alone. A pool of neuronal GAD67 co-localizes with GAD65 in the Golgi complex region and in many synapses. The two proteins also co-localize in the perinuclear region of some pancreatic beta-cells. GAD67 interacts with the NH2-terminal region of GAD65, even in the absence of palmitoylation of this region of GAD65. Taken together, our results indicate that GAD65-GAD67 association occurs in vivo and is required for the targeting of GAD67 to membranes.

Animals↗
PubMed2006

Antigen-presenting cells expressing glutamate decarboxylase 67 were identified as epithelial cells in glutamate decarboxylase 67-GFP knock-in mouse thymus.

Glutamate decarboxylase (GAD), which has two isoforms, GAD65, and GAD67, is responsible for synthesis of the major inhibitory neurotransmitter, gamma-aminobutyric acid. GAD is expressed predominantly in the central nervous system; recent reports suggest that GAD is also expressed in non-neuronal organs including the pancreas. In the pancreatic islets, GAD serves as one of the autoantigens in type I diabetes mellitus. Recent flow cytometric analyses have shown that a variety of self-antigens, including GAD, are ectopically transcribed and expressed in particular cell populations of the thymus, although consensus concerning the cellular phenotype has not been obtained. The aim of this study was to clarify the localization and cellular phenotype of GAD67-expressing cells in the thymus at a cellular level with a novel approach using GAD67-green fluorescent protein (GFP) knock-in mice, in which GFP is expressed specifically in GAD67-positive cells. GFP-positive cells were detected in the thymic medulla and were identified as epithelial cells by immunohistochemistry. Almost all GFP-positive cells were positive for major histocompatibility complex (MHC) class II antigen staining and were positive for both cytokeratin and Ulex Europaeus Agglutinin I, markers of medullary thymic epithelial cells, but were negative for CD11c, Gr-1, and CD45, markers of dendritic cells, macrophages, and B-lymphocytes, respectively.

Animals↗
PubMed1976

Studies of glutamate dehydrogenase. Regulation of glutamate dehydrogenase from Candida utilis by a pH and temperature-dependent conformational transition.

Glutamate dehydrogenase from Candida utilis undergoes a reversible conformational transition between an active and an inactive state at low pH AND low temperature. This conformational transition can also be followed by fluorescence measurements. The temperature-dependent equilibrium between the active and the inactive state is characterized by a transition temperature of 10.7 degrees C and a delta H value of 148 kcal/mol (620 kJ/mol). The temperature dependence of the enzymic activity above 15 degrees C yields an activation energy of 15 kcal/mol (63 kJ/mol), a larger value than that for the beef liver enzyme (9 kcal/mol; 38 kJ/mol). In contrast to the yeast enzyme the Arrhenius plot is linear and, therefore, the beef liver enzyme is not transformed into an inactive conformation at low temperatures. Sedimentation analysis shows that the inactivation of the Candida utilis enzyme is not caused by change in the quaternary structure. The pH dependence of the conformational transition at low pH measured by fluorescence change is characterized by a pK value of 7.01 for the enzyme in the absence and of 6.89 for the enzyme in the presence of 2-oxoglutarate with a Hill coefficient of 3.4 in both cases. Similar results are found when the pH dependence of the enzymic activity is analyzed. With the beef liver enzyme the same pK value is obtained but with a Hill coefficient of 1 indicating cooperativity only in the case of the Candida utilis enzyme. The best fit of the pH dependence of the rate constants of the fluorescence changes was obtained with pK values of 7.45 and 6.45 for the active and the inactive state respectively. In this model the lowest time constant which is obtained at the pH of the equilibrium was found to be 0.05 s-1. Preincubation experiments with the substrate 2-oxoglutarate but not with the coenzyme shift the equilibrium to the active conformation. The coenzyme obviously reduces the rate constant of the conformational transition. The sedimentation coefficient (SO20, w) and the molecular weight were found to be 11.0 S and 276 000, respectively. The enzyme molecule is built up by six polypeptide chains each having a molecular weight of 47 000.

Binding Sites↗
PubMed2005

Metabotropic glutamate receptors and neuroadaptation to antidepressants: imipramine-induced down-regulation of beta-adrenergic receptors in mice treated with metabotropic glutamate 2/3 receptor ligands.

Antidepressant drugs have a clinical latency that correlates with the development of neuroadaptive changes, including down-regulation of beta-adrenergic receptors in different brain regions. The identification of drugs that shorten this latency will have a great impact on the treatment of major depressive disorders. We report that the time required for the antidepressant imipramine to reduce the expression of beta-adrenergic receptors in the hippocampus is reduced by a co-administration with centrally active ligands of type 2/3 metabotropic glutamate (mGlu2/3) receptors. Daily treatment of mice with imipramine alone (10 mg/kg, i.p.) reduced the expression of beta-adrenergic receptors in the hippocampus after 21 days, but not at shorter times, as assessed by western blot analysis of beta1-adrenergic receptors and by the amount of specifically bound [3H]CGP-12177, a selective beta-adrenergic receptor ligand. Down-regulation of beta-adrenergic receptors occurred at shorter times (i.e. after 14 days) when imipramine was combined with low doses (0.5 mg/kg, i.p.) of the selective mGlu2/3 receptor agonist LY379268, or with the preferential mGlu2/3 receptor antagonist LY341495 (1 mg/kg, i.p.). Higher doses of LY379268 (2 mg/kg, i.p.) were inactive. This intriguing finding suggests that neuroadaptation to imipramine--at least as assessed by changes in the expression of beta1-adrenergic receptors--is influenced by drugs that interact with mGlu2/3 receptors and stimulates further research aimed at establishing whether any of these drugs can shorten the clinical latency of classical antidepressants.

Adaptation, Physiological↗
PubMed1994

Nucleotide sequence of a Porphyromonas gingivalis gene encoding a surface-associated glutamate dehydrogenase and construction of a glutamate dehydrogenase-deficient isogenic mutant.

The nucleotide sequence for a surface-associated protein (A. Joe, A. Yamamoto, and B. C. McBride, Infect. Immun. 61:3294-3303, 1993) of Porphyromonas gingivalis was determined. The structural gene comprises 1,338 bp and codes for a protein of 445 amino acids. The deduced molecular weight of the protein is 49,243. A data base search for homologous proteins revealed significant sequence similarity to the subunit protein of glutamate dehydrogenases (GDHs) isolated from various sources. This protein, which was previously labelled PgAg1, will now be called GDH. Recombinant GDH was purified to homogeneity, and native GDH was partially purified from P. gingivalis. Both preparations exhibited NAD-dependent GDH activity. Intact P. gingivalis and an extract of cell surface components also demonstrated NAD-dependent GDH activity. To help elucidate the role of this protein, an isogenic mutant of P. gingivalis lacking the GDH protein was generated by deletion disruption. Biological characterization of the mutant strain, P. gingivalis E51, demonstrated complete loss of GDH activity. Immunogold bead labelling of intact cells showed that GDH was no longer present on the surface of the bacterial cell. The GDH-negative mutant displayed impaired cell growth, as demonstrated by an increased generation time and an inability to grow to the same cell density as the parent.

Amino Acid Sequence↗
PubMed2005

Biosynthesis of poly-gamma-glutamic acid in plants: transient expression of poly-gamma-glutamate synthetase complex in tobacco leaves.

Transient expression of genes coding for the poly-gamma-glutamate (gammaPGA) synthetase system (pgs) was investigated in tobacco plants. Three genes of the pgs, pgsA, pgsB and pgsC, were separately placed under the control of the CaMV 35S promoter and introduced into tobacco leaves via Agrobacterium infection. Synthesized gammaPGA in plant tissues was detected immunologically with mouse anti-gammaPGA antiserum which specifically reacts with gammaPGA on a nitrocellulose membrane. Confirmation of gammaPGA biosynthesis in the transient expression analysis in tobacco tissue indicates that subunits of pgs complex were expressed and reassembled in a functional form.

Genes, Plant↗