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)

Recently we demonstrated that the unique stereoelectronic relationships inherent in the structure of plasmenylethanolamine facilitate membrane fusion, and we postulated the existence of a membrane fusion protein which could exploit the propensity of plasmenylethanolamine molecular species to adapt an inverted hexagonal phase [Glaser & Gross (1994) Biochemistry 33, 5805-5812]. We now report a cryptic membrane fusion activity in rabbit brain cytosol, which requires separation from an endogenous inhibitor to express its activity, and demonstrate that vesicle fusion catalyzed by this protein is highly selective for membrane vesicles containing plasmenylethanolamine. The cytosolic protein catalyzing membrane fusion activity was purified to apparent homogeneity by sequential column chromatographies, revealing a single 38-kDa protein band after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining. Automated Edman degradation demonstrated that the purified protein is an isoform of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which was confirmed by Western blot analysis utilizing polyclonal antibodies and by solution-state inactivation of membrane fusion activity by a monoclonal antibody directed against GAPDH. Both GTP-affinity and Mono Q chromatographies resolved GAPDH isoforms that catalyzed dehydrogenase activity from the GAPDH isoform that catalyzed membrane fusion activity. The purified fusion protein was calcium-independent, resistant to treatment with N-ethylmaleimide, and possessed an obligatory requirement for plasmenylethanolamine and cholesterol. High-resolution stopped-flow kinetic analysis of plasmenylethanolamine-facilitated membrane fusion demonstrated that one tetramer of the GAPDH isoform catalyzed one fusion event between two vesicles containing plasmenylethanolamine every millisecond (on average). Collectively, these results constitute the first description of a protein which can catalyze the fusion of vesicles at a rate which satisfies the mathematical constraints imposed by the observed rates of fusion of synaptic vesicles with the presynaptic membrane in vivo.
Biochemistry 1995 Sep 26
PMID:Rapid plasmenylethanolamine-selective fusion of membrane bilayers catalyzed by an isoform of glyceraldehyde-3-phosphate dehydrogenase: discrimination between glycolytic and fusogenic roles of individual isoforms. 754 60

In continuation of a project aimed at the structure-based design of drugs against sleeping sickness, analogs of 2'-deoxy-2'-(3-methoxybenzamido)adenosine (1) were synthesized and tested to establish structure-activity relationships for inhibiting glycosomal glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Compound 1 was recently designed using the NAD:GAPDH complexes of the human enzyme and that of Trypanosoma brucei, the causative agent of sleeping sickness. In an effort to exploit an extra hydrophobic domain due to Val 207 of the parasite enzyme, several new 2'-amido-2'-deoxyadenosines were synthesized. Some of them displayed an interesting improvement in inhibitory activity compared to 1. Carbocyclic or acyclic analogs showed marked loss of activity, illustrating the importance of the typical (C-2'-endo) puckering of the ribose moiety. We also describe the synthesis of a pair of compounds that combine the beneficial effects of a 2- and 8-substituted adenine moiety on potency with the beneficial effect of a 2'-amido moiety on selectivity. Unfortunately, in both cases, IC50 values demonstrate the incompatibility of these combined modifications. Finally, introduction of a hydrophobic 5'-amido group on 5'-deoxyadenosine enhances the inhibition of the protozoan enzyme significantly, although the gain in selectivity is mediocre.
J Med Chem 1995 Sep 15
PMID:Synthesis and structure-activity relationships of analogs of 2'-deoxy-2'-(3-methoxybenzamido)adenosine, a selective inhibitor of trypanosomal glycosomal glyceraldehyde-3-phosphate dehydrogenase. 756 15

Genes for glycolytic and Calvin-cycle glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of higher eukaryotes derive from ancient gene duplications which occurred in eubacterial genomes; both were transferred to the nucleus during the course of endosymbiosis. We have cloned cDNAs encoding chloroplast and cytosolic GAPDH from the early-branching photosynthetic protist Euglena gracilis and have determined the structure of its nuclear gene for cytosolic GAPDH. The gene contains four introns which possess unusual secondary structures, do not obey the GT-AG rule, and are flanked by 2- to 3-bp direct repeats. A gene phylogeny for these sequences in the context of eubacterial homologues indicates that euglenozoa, like higher eukaryotes, have obtained their GAPDH genes from eubacteria via endosymbiotic (organelle-to-nucleus) gene transfer. The data further suggest that the early-branching protists Giardia lamblia and Entamoeba histolytica--which lack mitochondria--and portions of the trypanosome lineage have acquired GAPDH genes from eubacterial donors which did not ultimately give rise to contemporary membrane-bound organelles. Evidence that "cryptic" (possibly ephemeral) endosymbioses during evolution may have entailed successful gene transfer is preserved in protist nuclear gene sequences.
Proc Natl Acad Sci U S A 1995 Sep 26
PMID:A nuclear gene of eubacterial origin in Euglena gracilis reflects cryptic endosymbioses during protist evolution. 756 85

Nitric oxide (NO), produced by vascular endothelial cells, mediates both physiological and pathological responses. Although the molecular targets responsible for NO-mediated endothelial cell injury are not known, one candidate is the glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In this study, we investigated the mechanism involved in NO-mediated GAPDH inhibition and found that S-nitrosoglutathione (GSNO) inhibited GAPDH activity in both purified enzyme preparations and endothelial cells. Furthermore, GSNO-mediated GAPDH inhibition occurred by modification of the active site cysteine residue in GAPDH, since increasing concentrations of the substrate, glyceraldehyde-3-phosphate, which interacts with the active site cysteine residue, protected GAPDH from inhibition by GSNO. Although under certain conditions both GSNO and the NO donor, sodium nitroprusside (SNP), led to the covalent NAD(+)-dependent modification of GAPDH, this putative ADP ribosylation was unlikely to be the primary mechanism for inhibition, since the stoichiometry was extremely low, and, in the case of GSNO, inhibition was completely reversed by thiol reagents. Furthermore, GSNO effectively S-nitrosylated GAPDH, and the extent of nitrosylation was linearly correlated with the degree of inhibition such that addition of 1 mole of NO per mole of GAPDH monomer was necessary to inhibit the enzyme. Consistent with this finding, GSNO-mediated GAPDH inhibition was reversible with low-molecular-weight thiols, and the reversal of inhibition correlated with the "denitrosylation" of GAPDH. These results suggest that endothelial GAPDH is a target for NO and that inhibition occurs principally by the reversible S-nitrosylation of the active site cysteine residue in GAPDH.
Am J Physiol 1995 Sep
PMID:S-nitrosoglutathione reversibly inhibits GAPDH by S-nitrosylation. 757 5

African trypanosomes are motile unicellular eukaryotes that can cause diseases such as sleeping sickness in humans and nagana in animals, debilitating millions of people and livestock. All members of the Trypanosomatidae family contain subpellicular microtubules cross-linked to each other and to the plasma membrane by unique trypanosomal microtubule-associated proteins (MAPs). These MAPs may serve as specific intracellular target sites for therapeutic attack against trypanosomiasis. A trypanosomal MAP (p52) copurifies with two glycosomal enzymes (aldolase and GAPDH) on phosphocellulose columns. Rats and mice vaccinated with antigen preparation p52 containing the glycosomal enzymes were protected against a potentially fatal Trypanosoma brucei infection. Sera of protected animals caused in vitro aggregation of trypanosomes, and immunoelectron microscopy of these aggregates located antibodies in the cytoplasm of the trypanosomes.
J Infect Dis 1995 Sep
PMID:Intracellular antigens (microtubule-associated protein copurified with glycosomal enzymes)--possible vaccines against trypanosomiasis. 765 80

S-nitro-N-acetyl-DL-penicillamine (SNAP), a nitric oxide (NO) donor, inactivated bovine glutathione peroxidase (GPx) in a dose- and time-dependent manner. The IC50 of SNAP for GPx was 2 microM at 1 h of incubation and was 20% of the IC50 for another thiol enzyme, glyceraldehyde-3-phosphate dehydrogenase, in which a specific cysteine residue is known to be nitrosylated. Incubation of the inactivated GPx with 5 mM dithiothreitol within 1 h restored about 50% of activity of the start of the SNAP incubation. A longer exposure to NO donors, however, irreversibly inactivated the enzyme. The similarity of the inactivation with SNAP and reactivation with dithiothreitol of GPx to that of glyceraldehyde-3-phosphate dehydrogenase, suggested that NO released from SNAP modified a cysteine-like essential residue on GPx. When U937 cells were incubated with 100 microM SNAP for 1 h, a significant decrease in GPx activity was observed although the change was less dramatic than that with the purified enzyme, and intracellular peroxide levels increased as judged by flow cytometric analysis using a peroxide-sensitive dye. Other major antioxidative enzymes, copper/zinc superoxide dismutase, manganese superoxide dismutase, and catalase, were not affected by SNAP, which suggested that the increased accumulation of peroxides in SNAP-treated cells was due to inhibition of GPx activity by NO. Moreover, stimulation with lipopolysaccharide significantly decreased intracellular GPx activity in RAW 264.7 cells, and this effect was blocked by NO synthase inhibitor N omega-methyl-L-arginine. This indicated that GPx was also inactivated by endogenous NO. This mechanism may at least in part explain the cytotoxic effects of NO on cells and NO-induced apoptotic cell death.
J Biol Chem 1995 Sep 08
PMID:Inactivation of glutathione peroxidase by nitric oxide. Implication for cytotoxicity. 767 30

Biochemical and molecular markers of inflammation, cell proliferation, and pulmonary fibrosis were studied in lungs and bronchoalveolar lavage preparations from Fischer 344 rats at time periods from 3 to 20 days after inhalation of two airborne concentrations (0.18 and 8.2 mg/m3 air) of chrysotile asbestos. Additional groups of animals were examined for lung histopathology and cell proliferation with an antibody to 5-bromo-2'-deoxyuridine after exposure to asbestos for 5 and 20 days and after 20 days of exposure followed by an additional 20 days in room air. Exposure to chrysotile at the higher concentration caused protracted increases in steady-state mRNA levels of manganese-containing superoxide dismutase and elevation in glyceraldehyde-3-phosphate dehydrogenase mRNA at 3 days, but levels of mRNAs encoding copper-zinc-containing superoxide dismutase, ornithine decarboxylase, and the proto-oncogene, c-jun were not statistically elevated from levels occurring in lung homogenates from sham control rats. Differential cell counts in bronchoalveolar lavage revealed an early infiltration of neutrophils that correlated with focal areas of increased cellularity and fibrosis in rat lungs at the higher concentrations of asbestos. However, elevations in lung hydroxyproline were not observed. Significant increases in epithelial cells of the bronchi, the interstitial compartment of the lung, and mesothelial cells incorporating 5-bromo-2'-deoxyuridine, an indication of DNA synthesis, were noted in the higher chrysotile group at 5 days, but labeling in all cell compartments was comparable with that occurring in sham controls at later time points. Indicators of inflammation, increased cell proliferation, and pulmonary fibrosis were not observed in the lungs of rats exposed to the lower concentration of chrysotile. Thus, results indicate that cellular and molecular markers of inflammation and proliferation in lung are dose-related and indicative of the histopathological development of asbestosis.
Am J Pathol 1995 Sep
PMID:Patterns of inflammation, cell proliferation, and related gene expression in lung after inhalation of chrysotile asbestos. 767 84

Nitric oxide (NO) has been suggested to act as a regulator of endogenous intracellular ADP-ribosylation, based on radiolabelling of proteins in tissue homogenates incubated with [32P]NAD and NO. After the NO-stimulated modification was replicated in a defined system containing only the purified acceptor protein, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), the hypothesis of NO-stimulation of an endogenous ADP-ribosyltransferase became moot. The NO-stimulated, NAD-dependent modification of GAPDH was recently characterized as covalent binding of the whole NAD molecule to the enzyme, not ADP-ribosylation. With this result, along with the knowledge that GAPDH is stoichiometrically S-nitrosylated, the role of NO in protein modification with NAD may be viewed as the conferring of an unexpected chemical reactivity upon GAPDH, possibly due to nitrosylation of a cysteine in the enzyme active site.
Mol Cell Biochem 1994 Sep
PMID:Nitric oxide and NAD-dependent protein modification. 789 64

The ability of compounds releasing nitric oxide (NO) to regulate glyceraldehyde-3-phosphate dehydrogenase (GraPDH) activity was analysed both in cell homogenates and in intact Dictyostelium discoideum. The time course of GraPDH inactivation in cell lysates by NO-releasing compounds suggests that two processes may be involved, one of which accounts for the majority of the inactivation and shows a close correlation with GraPDH ADP-ribosylation. Maximal ADP-ribosylation under these conditions exhibited a stoichiometry of about 0.4 mol ADP-ribose/mol enzyme tetramer. NO-mediated inhibition of GraPDH activity was attenuated if specific substrates, cofactors, or cysteine were added to cytosol preparations. Under such conditions, ADP-ribosylation of the enzyme was correspondingly reduced or negligible. Intact cells treated with NO-releasing compounds were shown to respond by rapidly decreasing their GraPDH activity. This inhibition was transient and, after a 10-min incubation, enzyme activity returned to the level seen in control cells. The time course of these in vivo changes correlated well with those of the NO-stimulated ADP-ribosylation of GraPDH also seen in intact cells. The basis underlying the NO-stimulated inhibition of GraPDH activity was investigated and found to reflect a decreased Vmax. No changes in either the Km of the enzyme for its substrates or its state of polymerization were observed.
Eur J Biochem 1994 Sep 01
PMID:Nitric oxide regulation of glyceraldehyde-3-phosphate dehydrogenase activity in Dictyostelium discoideum cells and lysates. 792 59

Exposure of endothelial cells (EC) to hypoxia results in the increased expression of a distinct set of proteins with molecular masses of 56, 47, 39, 36, and 34 kDa. Their induction appears to be unique to EC and the stress of decreased oxygen tension. To understand the mechanism(s) and significance of the up-regulation of these proteins we have identified the 36-kDa protein by limited amino-terminal amino acid sequencing. The 21-amino acid sequence from the bovine protein exhibited 90.5% identity with the human sequence of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Northern blot analysis showed that the time course and extent of EC GAPDH mRNA up-regulation correlated with the increase in 36-kDa protein synthesis. Nuclear runoff analysis demonstrated that this increase in GAPDH expression is regulated, in part, at the transcriptional level; however, the increase in the rate of transcription did not account for the entire mRNA accumulation, suggesting that GAPDH, like other hypoxia-regulated proteins, is posttranscriptionally regulated. Subcellular fractionation of hypoxic EC showed up-regulation of the 36-kDa protein in the cytoplasmic fraction and, to a lesser extent, in the nuclear fraction. The up-regulation of GAPDH in EC may be related to their relative hypoxia tolerance. Alternatively, the up-regulation of GAPDH in EC during hypoxia may be related to the potential nonglycolytic functions of this enzyme.
J Biol Chem 1994 Sep 30
PMID:Regulation of endothelial cell glyceraldehyde-3-phosphate dehydrogenase expression by hypoxia. 792 7


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