Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.2.1.13 (glyceraldehyde-3-phosphate dehydrogenase)
6,511 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

When rabbit muscle phosphoglycerate kinase (PGK; a 48-kDa monomeric protein) and glyceraldehyde-3-phosphate dehydrogenase (GraPDH; a 145-kDa homotetrameric protein) are present together in solution in the proportion of 1 mol PGK/1 mol GraPDH monomer (total protein 0.2-1.0 mg/ml), an 80--82-kDa protein species is observed by gel-penetration (dilution factor) method and by the conventional procedure of elution from a gel column. Individually, PGK and GraPDH do not exhibit any self association or dissociation in the concentration range employed. Electrophoresis of the 80-82-kDa peak eluted from the gel column shows a single protein band with mobility intermediate between those of GraPDH and PGK. In titration experiments by the gel-penetration method, plots of dilution factor of PGK (or GraPDH) activity versus GraPDH (or PGK) concentration shows two linear portions intersecting at approximately 1 mol GraPDH monomer/1 mol PGK. From the molecular-mass values and the titration experiments, it has been suggested that, in solution, these enzymes form a complex consisting of 1 molecule of PGK and one monomeric subunit of GraPDH (expected molecular mass 84 kDa). Its dissociation constant has been estimated to be equal to or less than 13 nM. The complex is dissociated in the presence of KCl or NADH, with approximately half dissociation at 0.1 M salt or 0.25 mM NADH. At 0.1 M KCl, the complex is completely dissociated by adding ATP, NADH or 3-phosphoglycerate. AMP, ADP, NAD+, glyceraldehyde-3-phosphate, phosphate ions and fructose-1,6-bisphosphate reverse the effect of KCl.
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PMID:Phosphoglycerate-kinase-glyceraldehyde-3-phosphate-dehydrogenase interaction. Molecular mass studies. 785 37

The crystal structure of holo-glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophile Thermotoga maritima was determined by Patterson search methods using the known structure of the Bacillus stearothermophilus enzyme. The structure was refined at a resolution of 2.5 A to an R-factor of 16.63% for 26289 reflections between 8.0 A an 2.5 A with F > 2 sigma(F). The crystallographic asymmetric unit contains two monomers related by approximate 2-fold symmetry and a tetramer is built up by crystallographic symmetry. The root-mean-square deviation of Ca positions of glyceraldehyde-3-phosphate dehydrogenase from T. maritima and B. stearothermophilus is 0.83 A in the NAD+ binding domains and smaller close to the cofactor. In contrast, the largest deviations in the catalytic domains are found at residues involved in coordination of sulphate ion SO4 339, which most likely marks the site of the attacking inorganic phosphate ion in catalysis. A large number of extra salt-bridges may be an important factor contributing to the high thermostability of this protein.
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PMID:The crystal structure of holo-glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic bacterium Thermotoga maritima at 2.5 A resolution. 787 72

Aldolase and glyceraldehyde-3-phosphate dehydrogenase from the extremely halophilic archaebacterium Haloarcula vallismortis are stable only in high concentrations of KCl present within the physiological environment. Data concerning the structural changes in the two enzymes as a result of lowering of salt concentration and changes in pH were obtained by monitoring the intrinsic protein fluorescence in the presence of quenchers. When the KCl concentrations were lowered below 2 M or in the presence of 6 M guanidine hydrochloride, the emission maximum shifted to a longer wavelength, indicating enhanced exposure of tryptophyl residues to the solvent. The spectral characteristics of the two proteins in guanidine hydrochloride and 0.4 M KCl were identical. However, these denatured states appear to be different than those observed after acid denaturation. Further perturbation of fluorescence was observed due to I-, and application of the Stern-Volmer law showed that the total fluorescence was available to the quenchers only in 0.4 M KCl solutions. The unfolding of proteins in 0.4 M KCl was a gradual process which was accompanied by a time-dependent loss in enzyme activity. The activity loss was complete within 30 min for aldolase whereas in the case of GAPDH nearly 3 h was required for the destruction of activity. For both enzymes, inactivation and protein denaturation were strongly correlated. The data on activity and thermostability measurements of the two enzymes in varying concentrations of KCl and potassium phosphate revealed that though both proteins are halophilic, the forces in the maintenance of their stability could be different.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Halophilic class I aldolase and glyceraldehyde-3-phosphate dehydrogenase: some salt-dependent structural features. 842 83

The crystal structure of holo D-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from the extreme thermophile Thermus aquaticus has been solved at 2.5 Angstroms resolution. To study the determinants of thermostability, we compare our structure to four other GAPDHs. Salt links, hydrogen bonds, buried surface area, packing density, surface to volume ratio, and stabilization of alpha-helices and beta-turns are analyzed. We find a strong correlation between thermostability and the number of hydrogen bonds between charged side chains and neutral partners. These charged-neutral hydrogen bonds provide electrostatic stabilization without the heavy desolvation penalty of salt links. The stability of thermophilic GAPDHs is also correlated with the number of intrasubunit salt links and total hydrogen bonds. Charged residues, therefore, play a dual role in stabilization by participating not only in salt links but also in hydrogen bonds with a neutral partner. Hydrophobic effects allow for discrimination between thermophiles and psychrophiles, but not within the GAPDH thermophiles. There is, however, an association between thermostability and decreasing enzyme surface to volume ratio. Finally, we describe several interactions present in both our GAPDH and a hyperthermophilic GAPDH that are absent in the less thermostable GAPDHs. These include a four-residue salt link network, a hydrogen bond near the active site, an intersubunit salt link, and several buried Ile residues.
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PMID:Determinants of enzyme thermostability observed in the molecular structure of Thermus aquaticus D-glyceraldehyde-3-phosphate dehydrogenase at 25 Angstroms Resolution. 861 63

We have recently shown that inhibition of endogenous Cu,Zn superoxide dismutase (SOD) by diethyldithiocarbamate (DDC) increased superoxide anion levels in isolated rabbit aortic rings, describing a useful experimental model to examine the effects of oxidative stress on the vessel wall. The present study examined the effects of oxidative stress on the steady-state mRNA levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.12). Aortic rings were incubated in physiological salt solution at 37 degrees C for up to 6 h. DDC (2 mM) decreased total SOD activity to < 5% of control levels and increased superoxide anion level ninefold. Steady-state mRNA levels of GAPDH were increased under comparable conditions. Although decreased biological activity of endothelium-derived nitric oxide was indicated by lower basal guanosine 3',5'-cyclic monophosphate levels in aortic rings treated with DDC compared with those in control rings (1.2 +/- 0.1 vs. 1.9 +/- 0.3 fmol/microgram protein, P < 0.05), neither endothelium denudation nor NG-nitro-L-arginine methyl ester had any effects on the steady-state mRNA levels of GAPDH. The cell.permeable iron chelator 1,10-phenanthroline completely prevented the increases in GAPDH mRNA levels induced by DDC. These results suggest that oxidative stress resulting from inhibition of endogenous Cu,Zn SOD causes induction of GAPDH gene expression and that the hydroxyl radical, produced through the iron-catalyzed Haber-Weiss reaction, is the intracellular reactive oxygen species responsible for the DDC-stimulated increase in GAPDH mRNA.
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PMID:Oxidative stress increases glyceraldehyde-3-phosphate dehydrogenase mRNA levels in isolated rabbit aorta. 876 37

Maize cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) is encoded by a small multi-gene family consisting of gpc1, gpc2, gpc3 and gpc4. GAPC3/4 protein is synthesized in roots during anoxic conditions and is known to be one of the 'anaerobic polypeptides'. We further analyzed the gpc gene family by isolating full-length cDNA clones of gpc2, gpc3, gpc4 and genomic clones of gpc2 and gpc4. The deduced amino acid sequence of GAPC4 has 99.4% identity with that of GAPC3 as compared to only 81% with either GAPC1 or GAPC2 amino acid sequence. Based on the deduced amino acid sequence identity we designated GAPC1 and GAPC2 as group I (97% identical) and GAPC3 and GAPC4 as group II (99.4% identical). As previously reported for gpc3, transcript levels were also induced for gpc4 by anaerobiosis. Neither heat shock, cold nor salt stress induced the expression of gpc3 or gpc4. In contrast, the transcript accumulation of gpc1 and gpc2 either remained constitutive or decreased in response to anoxia. The upstream regions of gpc2 and gpc4 contain typical eukaryotic promoter features with transcription start points at 76 and 68 bp upstream of their respective translation initiation sites. Transient expression analysis of gpc4 promoter-beta-glucuronidase (GUS) reporter gene constructs in bombarded maize suspension culture cells was used to examine the role of 5'-flanking sequence of gpc4. The gpc4 promoter (-1997 to +39 bp) was sufficient to induce GUS activity approximately three-fold in response to anaerobiosis. 5'-unidirectional deletion analysis revealed that the critical region of gpc4 required for its induced expression lies between -290 and -157. This region has reverse-oriented putative 'anaerobic response elements', G-box like sequences, and a GC motif similar to that previously defined as a regulatory element of maize adh1 and Arabidopsis adh, as well as the sequences found in other environmentally inducible genes. The relevance of these elements in conferring anaerobic induction of gpc4 gene expression is discussed.
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PMID:Molecular characterization and promoter analysis of the maize cytosolic glyceraldehyde 3-phosphate dehydrogenase gene family and its expression during anoxia. 903 63

Mutations have been introduced in the cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Bacillus stearothermophilus in order to convert its cofactor selectivity from a specificity towards NAD into a preference for NADP. In the B-S mutant, five mutations (L33T, T34G, D35G, L187A, P188S) were selected on the basis of a sequence alignment with NADP-dependent chloroplastic GAPDHs. In the D32G-S mutant, two of the five mutations mentioned above (L187A, P188S) have been used in combination with another one designed from electrostatic considerations (D32G). Both mutants exhibit a dual-cofactor selectivity at the advantage of either NAD (B-S) or NADP (D32G-S). In order to analyse the cofactor-binding site plasticity at the molecular level, crystal structures of these mutants have been solved, when complexed with either NAD+ (D32G-Sn, resolution 2.5 A, R = 13.9%; B-Sn, 2.45 A, 19.3%) or NADP+ (D32G-Sp, 2.2 A, 19.2%; B-Sp, 2.5 A, 14.4%). The four refined models are very similar to that of the wild-type GAPDH and as expected resemble more closely the holo form than the apo form. In the B-S mutant, the wild-type low affinity for NADP+ seems to be essentially retained because of repulsive electrostatic contacts between the extra 2'-phosphate and the unchanged carboxylate group of residue D32. Such an antideterminant effect is not well compensated by putative attractive interactions which had been expected to arise from the newly-introduced side-chains. In this mutant, recognition of NAD+ is slightly affected with respect to that known on the wild-type, because mutations only weakly destabilize hydrogen bonds and van der Waals contacts originally present in the natural enzyme. Thus, the B-S mutant does not mimic efficiently the chloroplastic GAPDHs, and long-range and/or second-layer effects, not easily predictable from visual inspection of three-dimensional structures, need to be taken into account for designing a true "chloroplastic-like" mutant of cytosolic GAPDH. In the case of the D32G-S mutant, the dissociation constants for NAD+ and NADP+ are practically reversed with respect to those of the wild-type. The strong alteration of the affinity for NAD+ obviously proceeds from the suppression of the two wild-type hydrogen bonds between the adenosine 2'- and 3'-hydroxyl positions and the D32 carboxylate group. As expected, the efficient recognition of NADP+ is partly promoted by the removal of intra-subunit electrostatic repulsion (D32G) and inter-subunit steric hindrance (L187A, P188S). Another interesting feature of the reshaped NADP+-binding site is provided by the local stabilization of the extra 2'-phosphate which forms a hydrogen bond with the side-chain hydroxyl group of the newly-introduced S188. When compared to the presently known natural NADP-binding clefts, this result clearly demonstrates that an absolute need for a salt-bridge involving the 2'-phosphate is not required to switch the cofactor selectivity from NAD to NADP. In fact, as it is the case in this mutant, only a moderately polar hydrogen bond can be sufficient to make the extra 2'-phosphate of NADP+ well recognized by a protein environment.
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PMID:A crystallographic comparison between mutated glyceraldehyde-3-phosphate dehydrogenases from Bacillus stearothermophilus complexed with either NAD+ or NADP+. 917 58

The aim of the study was to investigate whether the adrenal renin-angiotensin system plays an independent role in the regulation of mineralocorticoid biosynthesis in the adrenal gland and to explore the mechanisms of this action. Twelve-week-old male Sprague-Dawley rats were studied: 22 rats were maintained on a regular diet; 27 and 22 rats received a low salt diet with and without treatment, respectively, with the angiotensin II (Ang II) AT1-subtype receptor antagonist losartan (10 mg/kg per day). A fraction of each group of rats underwent bilateral nephrectomy (n = 12, 15, and 10, respectively) and was killed 48 hours later. In an additional group of 24 (12 intact and 12 nephrectomized) rats, the effects of the Ang II AT2-subtype receptor antagonist PD123319 were investigated. In intact rats, plasma renin activity (PRA) and adrenal renin activity and expression were progressively raised by salt restriction and losartan, whereas aldosterone synthase mRNA and plasma aldosterone (PA) levels were increased by salt restriction and reduced by losartan. Forty-eight hours after nephrectomy, PRA fell to undetectable levels; in contrast, adrenal renin expression, assessed by semiquantitative reverse-transcriptase polymerase chain reaction (using GAPDH as a standard for gene expression), showed an 18-fold increase and was further increased after salt restriction and losartan (all P < .05). Also, adrenal renin activity was raised after nephrectomy and further increased after salt restriction (P < .05) and losartan. Cytochrome P450 aldosterone synthase expression in the adrenal cortex was stimulated by nephrectomy alone and by nephrectomy combined with low salt intake (P < .05), with consequent increases in PA concentrations. In losartan-treated salt-restricted nephrectomized rats, cytochrome P450 aldosterone synthase expression (P < .05 versus nephrectomy alone and nephrectomy plus salt restriction) and PA concentrations were diminished (P < .05) in spite of the observed increases of adrenal renin expression. The AT2-receptor antagonism did not significantly affect PRA, adrenal renin, and aldosterone biosynthesis and production in either intact or nephrectomized salt-restricted rats. These results demonstrate that the adrenal renin-angiotensin system plays an independent role in the regulation of mineralocorticoid biosynthesis in vivo. This action is mediated primarily via the Ang II AT1-subtype receptors.
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PMID:Role of tissue renin in the regulation of aldosterone biosynthesis in the adrenal cortex of nephrectomized rats. 935 60

The structure of the enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from glycosomes of the parasite Trypanosoma cruzi, causative agent of Chagas' disease, is reported. The final model at 2.8 A includes the bound cofactor NAD+ and 90 water molecules per monomer and resulted in an Rfactor of 20.1%, Rfree = 22.3%, with good geometry indicators. The structure has no ions bound at the active site resulting in a large change in the side chain conformation of Arg249 which as a consequence forms a salt bridge to Asp210 in the present structure. We propose that this conformational change could be important for the reaction mechanism and possibly a common feature of many GAPDH structures. Comparison with the human enzyme indicates that interfering with this salt bridge could be a new approach to specific inhibitor design, as the equivalent to Asp210 is a leucine in the mammalian enzymes.
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PMID:Trypanosoma cruzi glycosomal glyceraldehyde-3-phosphate dehydrogenase: structure, catalytic mechanism and targeted inhibitor design. 958 Jan 89

Rate studies using phosphoglycerate kinase (PGK)--glyceraldehyde-3-phosphate dehydrogenase (GPDH) enzyme pair have been carried out to distinguish between the two mechanisms of intermediate metabolite transfer, namely diffusion through the solvent versus "substrate channelling" within an enzyme-enzyme complex. A procedure has been described for the assay of the rates of PGK-catalysed and the PGK-GPDH coupled reactions at high (saturating) GPDH concentration. With PGKs of rabbit muscle and yeast, the coupled reaction proceeded faster than the PGK-catalysed reaction. At a high salt concentration (0.5 M KCl), where a PGK-GPDH complex is known to dissociate, the two reactions proceeded at almost equal rates. At fixed PGK concentration, the rate of the coupled reaction at high (saturating) GPDH concentration varied with the nature (biological origin) of the latter enzyme. In the presence of 0.5 M KCl, the saturating rate values with different GPDHs were almost equal. The PGK-catalysed reaction exhibited typical Michaelian behaviour on varying the substrate concentrations (linear double reciprocal plots). The Km values for 3-PGA (0.51 mM) and ATP (0.40 mM) were independent of the concentration of the second substrate. The double reciprocal plots for the coupled reaction showed downward curvature, i.e. activation at higher substrate concentrations. The ratio of the rate of the coupled reaction: the rate of the PGK catalysed reaction was found to be a function of the nature of PGK, nature of GPDH, nature of buffer, pH, salt concentration and substrate concentrations. The ratio varied between close to unity at low substrate concentrations, to three when the Vmax values of the two reactions were compared. At low substrate concentrations, the rate of the coupled reaction became independent of the nature of GPDH. It has been suggested that in the PGK-GPDH pair, the intermediate metabolite (BPG) is transferred directly from one enzyme to the other within an enzyme-enzyme complex, except at high salt or low substrate concentrations. Under the latter conditions, data were consistent with metabolite transfer by diffusion. Implications of these results for coupled enzyme assays have been discussed.
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PMID:Phosphoglycerate kinase--glyceraldehyde-3-phosphate dehydrogenase interaction: reaction rate studies. 1054 68


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