Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Glutamate decarboxylase from barley embryos and roots. General properties and the occurrence of three enzymic forms.

ABSTRACT: Glutamate decarboxylase in extracts of barley has a Km value for L-glutamate of 22 mM and is activated by the addition of pyridoxal phosphate by up to 3.5 times. Sucrose-density-gradient experiments indicate the presence of two enzyme forms with molecular weights 256000 and 120000. The lower-molecular-weight form appears to be relatively inactive and spontaneously associates to the higher-molecular-weight form on storage. The enzyme is inhibited by thiol reagents and the distribution of activity on density gradients is altered in favour of the lower-molecular-weight form by the presence of 2-mercaptoethanol. After removal of the 2-mercaptoethanol spontaneous association to the higher-molecular-weight form occurs. The presence of oxygen in the extraction buffer and in the water during imbibition leads to a relative increase in the higher-molecular-weight form compared with situations where oxygen is excluded. In contrast, glutamate decarboxylase in extracts of 3-day-old barley roots has a Km value for L-glutamate of 3.1 mM and is activated up to 10% by addition of pyridoxal phosphate. The root enzyme occurs as a single species with molecular weight 310000 and this is unaffected by 2-mercaptoethanol although thiol reagents do act as weak inhibitors. The molecular weight is also unaffected by the presence or absence of oxygen in the extraction buffers.

SUBMITTER: Inatomi K

PROVIDER: S-EPMC1165475 | biostudies-other | 1975 Jun

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Three Distinct Glutamate Decarboxylase Genes in Vertebrates.

Project description:Gamma-aminobutyric acid (GABA) is a widely conserved signaling molecule that in animals has been adapted as a neurotransmitter. GABA is synthesized from the amino acid glutamate by the action of glutamate decarboxylases (GADs). Two vertebrate genes, GAD1 and GAD2, encode distinct GAD proteins: GAD67 and GAD65, respectively. We have identified a third vertebrate GAD gene, GAD3. This gene is conserved in fishes as well as tetrapods. We analyzed protein sequence, gene structure, synteny, and phylogenetics to identify GAD3 as a homolog of GAD1 and GAD2. Interestingly, we found that GAD3 was lost in the hominid lineage. Because of the importance of GABA as a neurotransmitter, GAD3 may play important roles in vertebrate nervous systems.

| S-EPMC4962313 | biostudies-literature

Characterization of three kinetically distinct forms of glutamate decarboxylase from pig brain.

Project description:Pig brain contains three forms of glutamate decarboxylase with pI values of 5.3, 5.5 and 5.8, referred to as the alpha-, beta- and gamma-forms respectively. These forms were purified and kinetically characterized. The major synaptic form of glutamate decarboxylase (the beta-form) migrated as a single band on electrophoresis in sodium dodecyl sulphate/polyacrylamide gels with an apparent Mr of 60 000. Sodium dodecyl sulphate/polyacrylamide gel electrophoresis followed by immunoblotting with an affinity-purified antibody to the enzyme indicated a subunit Mr of 60 000 for the alpha- and gamma-forms as well. An extensive kinetic analysis, aided by an integrated equation that describes the inactivation and re-activation cycle of the enzyme, revealed that the three forms of the enzyme differ markedly in kinetic properties. The Km values for L-glutamate were 0.17, 0.45 and 1.24 mM respectively for the alpha-, beta- and gamma-forms. The Ki for 4-aminobutyrate, the first-order rate constants for inactivation by L-glutamate and 4-aminobutyrate, the rate constant for re-activation of the apoenzyme by pyridoxal 5'-phosphate and the dissociation constant for pyridoxal 5'-phosphate also differed in a similar way among the three forms; the values were in the order alpha-form less than beta-form less than gamma-form.

| S-EPMC1152805 | biostudies-other

Purification and general properties of glutamate decarboxylase from Clostridium perfringens.

Project description:1. A seven-step procedure for preparing highly purified glutamate decarboxylase from Clostridium perfringens is described. 2. The homogeneity of the pure enzyme was established by sucrose-density-gradient centrifugation and starch-gel electrophoresis. 3. The isoelectric point of the pure enzyme is about pH4.5 and the molecular weight is 290000. 4. The pH optimum for activity is 4.7. The pure enzyme is specific for l-glutamate; beta-hydroxyglutamate is decarboxylated at a lower rate. 5. Evidence is presented that each mol of enzyme contains 2mol of firmly bound pyridoxal 5-phosphate. 6. Resolution does not occur at acid pH; by dilution with neutral or alkaline buffers the enzyme is inactivated and the coenzyme is released. 7. Reconstitution of active enzyme was obtained by protecting the apoenzyme with thiol compounds.

| S-EPMC1179089 | biostudies-other

An examination of aspartate decarboxylase and glutamate decarboxylase activity in mosquitoes.

Project description:A major pathway of beta-alanine synthesis in insects is through the alpha-decarboxylation of aspartate, but the enzyme involved in the decarboxylation of aspartate has not been clearly defined in mosquitoes and characterized in any insect species. In this study, we expressed two putative mosquito glutamate decarboxylase-like enzymes of mosquitoes and critically analyzed their substrate specificity and biochemical properties. Our results provide clear biochemical evidence establishing that one of them is an aspartate decarboxylase and the other is a glutamate decarboxylase. The mosquito aspartate decarboxylase functions exclusively on the production of beta-alanine with no activity with glutamate. Likewise the mosquito glutamate decarboxylase is highly specific to glutamate with essentially no activity with aspartate. Although insect aspartate decarboxylase shares high sequence identity with glutamate decarboxylase, we are able to closely predict aspartate decarboxylase from glutamate decarboxylase based on the difference of their active site residues.

| S-EPMC2913154 | biostudies-literature

RNA Sequencing Analysis of MC38 tumors with Glutamate decarboxylase 1 Knockdown

Project description:Various types of cancers, including colorectal cancer, have shown an unusual dependence on glutamine. Glutamine can undergo an alternate fate in neurons: conversion into a non-proteinogenic amino acid gamma-aminobutyric acid (GABA) by the glutamic acid decarboxylase (GAD), including two family members GAD1 and GAD2. Some studies have shown that GAD1 expression is dysregulated in certain cancer types, but the effect of GAD1 on the tumorigenic process remain largely unknown. Thus, we use mouse colon adenocarcinoma MC38 tumor samples to perform RNA-sequencing (RNA-seq) assay. The goal of this study is to investigate GAD1-mediated transcriptome changes within tumors.

2021-02-02 | GSE165889 | GEO

Mercury toxicity in barley roots

Project description:The effects of mercury (HgCl2) on barley (Hordeum vulgare L.) growth, physiological traits and gene expression profiles were studied. The shoot to root ratio was decreased in the two levels of HgCl2 (500 and 1000 μM) assayed, which was related primarily with decreases in shoot dry weight. Moreover stomatal conductance was limited and leaf carbon isotope discrimination decreased. Therefore water uptake limitations seem to be an important component of barley responses to HgCl2. Evidences for decreased stomatal conductance and water uptake limitations were further confirmed by the over expression of ABA related transcripts and down regulation of an aquaporin in roots. Root dry weight was only affected at 1000 μM HgCl2 and root browning was observed, while several transcripts for lignin biosynthesis were up regulated in HgCl2. Microarray analysis further revealed that growth inhibition in HgCl2 was related to increased expression of genes participating in ethylene biosynthesis and down regulation of several genes participating in DNA synthesis, chromatin structure and cell division, cell wall degradation and modification, oxidative pentose phosphate cycle and nitrogen metabolism pathway. Genes involved in detoxification and defence mechanisms were up regulated including several cytochrome P450s, glucosyltransferases and glutathione-s-transferases and amino acid metabolism participatory genes. It is concluded that barley plants survive in the presence of HgCl2 through several mechanisms that include water uptake limitations, shoot and root growth regulation, increased expression of genes involved in the biosynthesis of several plant protection secondary metabolites and finally through detoxification.

2009-05-13 | GSE15295 | GEO

Paenibacillus polymyxa invades plant roots and forms biofilms.

Project description:Paenibacillus polymyxa is a plant growth-promoting rhizobacterium with a broad host range, but so far the use of this organism as a biocontrol agent has not been very efficient. In previous work we showed that this bacterium protects Arabidopsis thaliana against pathogens and abiotic stress (S. Timmusk and E. G. H. Wagner, Mol. Plant-Microbe Interact. 12:951-959, 1999; S. Timmusk, P. van West, N. A. R. Gow, and E. G. H. Wagner, p. 1-28, in Mechanism of action of the plant growth promoting bacterium Paenibacillus polymyxa, 2003). Here, we studied colonization of plant roots by a natural isolate of P. polymyxa which had been tagged with a plasmid-borne gfp gene. Fluorescence microscopy and electron scanning microscopy indicated that the bacteria colonized predominantly the root tip, where they formed biofilms. Accumulation of bacteria was observed in the intercellular spaces outside the vascular cylinder. Systemic spreading did not occur, as indicated by the absence of bacteria in aerial tissues. Studies were performed in both a gnotobiotic system and a soil system. The fact that similar observations were made in both systems suggests that colonization by this bacterium can be studied in a more defined system. Problems associated with green fluorescent protein tagging of natural isolates and deleterious effects of the plant growth-promoting bacteria are discussed.

| S-EPMC1287669 | biostudies-literature

Crystal structure and functional analysis of Escherichia coli glutamate decarboxylase.

Project description:Glutamate decarboxylase is a vitamin B6-dependent enzyme, which catalyses the decarboxylation of glutamate to gamma-aminobutyrate. In Escherichia coli, expression of glutamate decarboxylase (GadB), a 330 kDa hexamer, is induced to maintain the physiological pH under acidic conditions, like those of the passage through the stomach en route to the intestine. GadB, together with the antiporter GadC, constitutes the gad acid resistance system, which confers the ability for bacterial survival for at least 2 h in a strongly acidic environment. GadB undergoes a pH-dependent conformational change and exhibits an activity optimum at low pH. We determined the crystal structures of GadB at acidic and neutral pH. They reveal the molecular details of the conformational change and the structural basis for the acidic pH optimum. We demonstrate that the enzyme is localized exclusively in the cytoplasm at neutral pH, but is recruited to the membrane when the pH falls. We show by structure-based site-directed mutagenesis that the triple helix bundle formed by the N-termini of the protein at acidic pH is the major determinant for this behaviour.

| S-EPMC175793 | biostudies-literature

Parallel Strategy Increases the Thermostability and Activity of Glutamate Decarboxylase.

Project description:Glutamate decarboxylase (GAD; EC 4.1.1.15) is a unique pyridoxal 5-phosphate (PLP)-dependent enzyme that specifically catalyzes the decarboxylation of L-glutamic acid to produce ?-aminobutyric acid (GABA), which exhibits several well-known physiological functions. However, glutamate decarboxylase from different sources has the common problem of poor thermostability that affects its application in industry. In this study, a parallel strategy comprising sequential analysis and free energy calculation was applied to identify critical amino acid sites affecting thermostability of GAD and select proper mutation contributing to improve structure rigidity of the enzyme. Two mutant enzymes, D203E and S325A, with higher thermostability were obtained, and their semi-inactivation temperature (T5015) values were 2.3 °C and 1.4 °C higher than the corresponding value of the wild-type enzyme (WT), respectively. Moreover, the mutant, S325A, exhibited enhanced activity compared to the wild type, with a 1.67-fold increase. The parallel strategy presented in this work proved to be an efficient tool for the reinforcement of protein thermostability.

| S-EPMC7037157 | biostudies-literature

Occurrence of twin embryos in the eastern bluebird.

Project description:We report the first record of presumed twinning in eastern bluebird (Sialia sialis) and provide a review of previously reported twinning events in wild birds. A nest containing twin eastern bluebird nestlings was monitored in 2013 in central Pennsylvania and reported to the Cornell Lab of Ornithology's NestWatch program, a national program where volunteers submit data on wild nesting birds. A presumed double-yolked egg of a free-living eastern bluebird pair hatched successfully, and twin nestlings lived for 11 days in a nest box shared by three siblings. Due to the rarity of twinning in wild birds, engaging the public to monitor large numbers of nests is the most likely approach to documenting twinning in wild populations, and citizen science provides the infrastructure for individuals to share observations.

| S-EPMC3961165 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data