Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.9 (glucose-6-phosphatase)
3,081 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Apolipoprotein B (apoB), a protein containing several hydrophobic beta-sheet structures, is essential for the assembly of triglyceride-rich lipoproteins. Previously, we found that only a fraction of de novo synthesized apoB is secreted; the remainder is retained in the endoplasmic reticulum where it is degraded. To understand the basis for these observations, translocation, the first step in the secretory pathway, was examined. Translocation of apoB was determined by its sensitivity to degradation by the exogenous protease, trypsin. In rough microsomes, about half of the apoB was degraded by trypsin. In contrast, in Golgi fractions little (if any) apoB was accessible to trypsin. Essentially all of the apoB that was degraded was membrane bound. Monoclonal IgGs against either the N-terminal or C-terminal halves of apoB were bound to magnetic beads and used to immunoisolate microsomes. In contrast to the specific ability of the IgGs against apoB to isolate microsomes, little or no microsomes were isolated using nonimmune IgG and IgG against albumin. Since microsomes remained intact and oriented right-side out as demonstrated by the inability of trypsin both to degrade albumin and to affect the capacity of the intralumenal enzyme glucose-6-phosphatase to dephosphorylate mannose 6-phosphate, the data suggest that a pool of apoB is exposed on the cytoplasmic surface of the endoplasmic reticulum membrane. To determine if the trypsin-accessible pool of apoB is a transient form, pulse-chase experiments were performed. The results show that the percent of apoB that was trypsin accessible increased during the first 20 min of the chase, suggesting that during this time the trypsin-accessible pool of apoB is not translocated (it does not become trypsin insensitive). Thus, in two in vivo models (cultured cells and rat liver) translocation of apoB is not quantitative. We propose that apoB translocation across the endoplasmic reticulum determines its entry into two functionally distinct pools. The intralumenal trypsin-insensitive pool participates in the assembly of very low density lipoprotein; the trypsin-accessible nontranslocated cytoplasmic pool is shunted into a degradative pathway. Regulated translocation of apoB may provide a unique mechanism with which to determine the rate of very low density lipoprotein assembly/secretion.
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PMID:Apolipoprotein B is both integrated into and translocated across the endoplasmic reticulum membrane. Evidence for two functionally distinct pools. 216 29

It was known in the 1950s that hepatic microsomal glucose-6-phosphatase plays an important role in the regulation of blood glucose levels. All attempts since then to purify a single polypeptide with glucose-6-phosphatase activity have failed. Until recently, virtually nothing was known about the molecular basis of glucose-6-phosphatase or its regulation. Recent studies of the type 1 glycogen storage diseases, which are human genetic deficiencies that result in impaired glucose-6-phosphatase activity, have greatly increased our understanding of glucose-6-phosphatase. Glucose-6-phosphatase has been shown to comprise at least five different polypeptides, the catalytic subunit of glucose-6-phosphatase with its active site situated in the lumen of the endoplasmic reticulum; a regulatory Ca2+ binding protein; and three transport proteins, T1, T2, and T3, which respectively allow glucose-6-phosphate, phosphate, and glucose to cross the endoplasmic reticulum membrane. Purified glucose-6-phosphatase proteins, immunospecific antibodies, and improved assay techniques have led to the diagnosis of a variety of new type 1 glycogen storage diseases. Recent studies of the type 1 glycogen storage diseases have led to a much greater understanding of the role and regulation of each of the glucose-6-phosphatase proteins.
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PMID:Molecular pathology of glucose-6-phosphatase. 216 25

We describe a novel technique for the histochemical and cytochemical demonstration of glucose-6-phosphatase activity. In this method, lead is replaced by cobalt. After activity of glucose-6-phosphatase, cobalt phosphate Co3(PO4)2 is formed, and in the presence of ammonium sulfide (NH4)2S, the precipitate is transformed into a sulfide that fixes osmium and provides good electron density. Glucose-6-phosphatase activity was determined mostly in rat kidney cells, but controls were also performed in liver cells. A strong reaction was seen in proximal tubule cells, but the reaction was weak in distal convoluted tubule cells. This technique showed the same endoplasmic reticulum (ER) organization in proximal and distal nephron as that seen with the osmium impregnation technique. In collecting tubules, intercalated cells had irregular reactivity, while principal cells had none. Our results indicate that the cobalt technique is valid, reliable, and sensitive enough to detect low glucose-6-phosphatase activity. Moreover, the technique can be used with 1-mm-thick specimens and obviates the need for use of frozen tissue sections.
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PMID:A new method based on cobalt for histochemical and cytochemical demonstration of glucose-6-phosphatase activity. 216 94

Microsomes from ventral prostate of 24-h castrated rats contain a single set of tissue-specific high-affinity, low-capacity androgen binding sites. These sites are indigenous to the endoplasmic reticulum, as shown by purification procedures associated with marker enzymes and electron microscopic analyses. When prostatic microsomal membranes are separated from plasma membranes using the nuclear or the mitochondrial pellets as the source of fractionation in sucrose gradients, the androgen binding activity is selectively associated with fractions rich in rough endoplasmic reticulum and ribosomes. Eighty-four percent of the total content of Na+/K+ adenosine triphosphatase (ATPase) and only 27% of the total binding capacity were concentrated in fractions rich in smooth-surfaced vesicular membranes, when nuclear suspensions constituted the membrane source. In contrast, the region of the same gradient when enriched in rough endoplasmic reticulum and deficient in plasma membrane content contained 73% of the androgen-binding capacity and only 14% of the ATPase. For fractions collected using mitochondrial suspensions as starting material, the ratio (total glucose-6-phosphatase/total binding capacity) was closer to 1.0 than similar ratios of ATPase/binding capacity, indicating co-sedimentation of binding sites with microsomal membranes and not with plasma membranes. Na+/K+ ATPase, but not 5' nucleotidase, is a valid plasma membrane marker for ventral prostate. Microsomal androgen receptors may constitute a new level of regulation of androgen action in target cells.
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PMID:Association of androgen binding sites with the endoplasmic reticulum of rat ventral prostate. 233 29

Aging is accompanied by a myriad of changes in cell structure, function, and composition. The fact that much of the information concerning age-related alterations in cellular morphology is qualitative precludes meaningful correlations with biochemical changes in order to enhance data interpretation. The mammalian liver has been subjected to both qualitative and quantitative evaluations of hepatocyte structure as a function of aging, i.e., development, maturation, and senescence. Although these data are characterized by considerable variability and, in some instances, blatant contradictions, there exists sufficient agreement in several parameters to permit a consensus in the inbred rat model. Certainly the volume of individual hepatocytes increases with age, and many of the organelle compartments reflect this change. While old rats exhibit a high incidence of polyploidy, there is no definitive evidence to demonstrate a concomitant increase in the binuclear hepatocyte index. Several specific hepatocellular organelles undergo changes in their relative volume or surface area that appear to correlate with functional alterations. The volume density of the lysosomal compartment enlarges significantly during senescence and is accompanied by increased activities of several constituent hydrolases. The hepatic concentration of smooth-surfaced endoplasmic reticulum declines markedly with aging, as does the yield of liver microsomes and the activities of several microsomal enzymes, e.g., mono-oxygenases and glucose-6-phosphatase. However, the responses of the majority of hepatocyte organelles to aging is varied and inconsistent based on the limited data currently available.
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PMID:Hepatocyte fine structure during maturation and senescence. 240 86

The lectin wheat germ agglutinin (WGA) conjugated to horseradish peroxidase (HRP) was employed to study the endocytic and exocytic pathways of the secretory process in neurons and the potential for trans-synaptic transfer of molecules within the CNS. WGA-HRP binds to surface membrane oligosaccharides and enters cells by adsorptive endocytosis. The lectin conjugate was administered intranasally or into the cerebral ventricles of mice; postinjection survival times ranged from 5 minutes to 6 days. Due to binding of the lectin to ependymal cells subsequent to an intraventricular injection, only select populations of neurons (i.e., hippocampal formation; paraventricular nuclei; midbrain raphe; VI, X, XII motor nuclei; among others) were exposed extracellularly to WGA-HRP and became labeled by retrograde axoplasmic transport from axon terminals or by direct cell body/dendritic uptake. WGA-HRP delivered intranasally was endocytosed by first-order olfactory neurons and transported by anterograde axoplasmic flow to the terminal field within the glomerular layer of the main olfactory bulb; eventually perikarya of the mitral cell layer were labeled, presumably by anterograde trans-synaptic transfer of the lectin conjugate. In the variety of neurons analyzed ultrastructurally following exposure to WGA-HRP, the proposed sequence of intracellular pathways through which peroxidase reaction product was traced over time was: cell surface membrane----endocytic structures----endosomes (presecondary lysosomes)----transfer vesicles----transmost Golgi saccule----vesicles, vacuoles, and/or dense core granules. WGA-HRP also labeled vesicles and tubules that were channeled to and/or derived from spherical endosomes, dense bodies, and multivesicular bodies. The peroxidase-positive, membrane-delimited products of the trans Golgi saccule contributed to anterograde axonal transport vectors and accumulated within axon terminals. A second contribution to these vectors was provided by peroxidase-labeled tubules and dense bodies believed to represent components of the lysosomal compartment. Profiles of the axonal reticulum comparable to those that stained cytochemically for glucose-6-phosphatase activity, a marker for the endoplasmic reticulum, were not associated with the transport of WGA-HRP. Trans-synaptic transfer of WGA-HRP from primary olfactory neurons to postsynaptic cells in the olfactory bulb was reflected in peroxidase-positive endocytic vesicles, endosomes, dense bodies, and the trans Golgi saccule.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Endocytic and exocytic pathways of the neuronal secretory process and trans-synaptic transfer of wheat germ agglutinin-horseradish peroxidase in vivo. 241 83

Inositol 1,4,5-trisphosphate [Ins(1,4,5)P3], arising from hydrolysis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2], is proposed as the link between membrane-receptor activation and mobilization of Ca2+ from intracellular sites in hormone-secreting cells. The location of Ins(1,4,5)P3-sensitive membranes was investigated in cultured neonatal beta-cells. Membranes were obtained after lysis of cells attached to positively charged Sephadex. After lysis the presence of the enzyme markers 5'-nucleotidase, glucose-6-phosphatase, NADH-cytochrome c reductase, UDP-galactosyltransferase and succinate dehydrogenase indicated the mixed nature of the preparation. After sonication, however, UDP-galactosyltransferase and succinate dehydrogenase activities were undetectable, but 4.8% of total cellular glucose-6-phosphatase and 3.4% of total cellular NADH-cytochrome c reductase remained with 5'-nucleotidase in the preparation, indicating endoplasmic-reticulum association. ATP-dependent 45Ca2+ accumulation was shown in this preparation (410 +/- 24 pmol/mg of protein at 150 nM free Ca2+) and was inhibited by vanadate (100 microM). Ca2+ release was effected by Ins(1,4,5)P3, with half-maximal release at 0.5 +/- 0.14 microM-Ins(1,4,5)P3, t1/2 11.2 +/- 1.1 s. GTP- and guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG)-promoted release of 45Ca2+ was demonstrated in this preparation, but the kinetics of release (half-maximal Ca2+ release at 5.4 +/- 0.7 microM, with t1/2 77.3 +/- 6.9 s, and at 51.1 +/- 4.2 microM, with t1/2 19.0 +/- 2.2 s, for GTP and p[NH]ppG respectively), and the ability of neomycin sulphate to block p[NH]ppG-induced release only, are indicative of separate release mechanisms after treatment with these agents. A close association between plasma membrane and elements of the endoplasmic reticulum is indicated in this model, providing a possible mechanism for local alterations in free Ca2+ in the sub-plasma-membrane region.
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PMID:GTP- and inositol 1,4,5-trisphosphate-induced release of 45Ca2+ from a membrane store co-localized with pancreatic-islet-cell plasma membrane. 245 19

In order to study the characteristics of the intracellular Ca store of mast cells, organelles of rat peritoneal mast cells were fractionated. The binding of 45Ca was at its peak in the fractions where the highest activity of glucose-6-phosphatase, the marker enzyme for the endoplasmic reticulum (ER), was measured. The ER-rich fraction exhibited an ATP-dependent uptake of 45Ca and this uptake was inhibited by pretreatment with ATPase inhibitors such as LaCl3 or Na3VO4. When inositol 1,4,5-trisphosphate (IP3) was added to a medium containing the 45Ca-loaded ER fraction, it caused a dose-dependent release of 45Ca at concentrations higher than 0.5 microM, while inositol 1-monophosphate and inositol 1,4-bisphosphate were not effective even at higher concentrations. The results of a binding assay using 3H-labeled IP3 indicated that there exist two kinds of IP3 binding site in the ER: one is of high affinity but low capacity while the other is of low affinity and high capacity. IP3-induced 45Ca release was dose-dependently inhibited by pretreatment with c-AMP. The present study supports the assumption that the intracellular Ca store associated with histamine release from the mast cell is the ER.
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PMID:Ca uptake and Ca releasing properties of the endoplasmic reticulum in rat peritoneal mast cells. 246 54

AtT-20 cells, which were derived from a murine pituitary tumor and produce ACTH, have until now been considered to originate from pituitary corticotrophs. Here we show that AtT-20 cells constitutively express several neuronal features. First, AtT-20 cells develop cytoplasmic processes whose fine structure is essentially identical to that of neurites and neuronal growth cones. These growth cones (i) are characterized by an extensive membranous reticulum which is derived from the endoplasmic reticulum (ER) since it contains immunoglobulin heavy chain binding protein, protein disulfide isomerase and glucose-6-phosphatase; (ii) are a major site of endocytosis; (iii) form cell-to-cell contacts resembling immature synapses. Second, AtT-20 cells, in contrast to pituitary corticotrophs, contain neurofilaments and express all three neurofilament polypeptides. They also contain the high molecular weight form of microtubule-associated protein 2 and tau protein. Third, AtT-20 cells express the neuron-specific phosphoprotein synapsin I which accumulates in the growth cones prior to contacts forming between growth cones and cells. Our results show that AtT-20 cells exhibit several properties of peptidergic neuronal cells and that the constitutive expression of a variety of these properties is compatible with continuous cell division.
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PMID:Morphological and biochemical evidence showing neuronal properties in AtT-20 cells and their growth cones. 250 49

We previously reported that phenylmethylsulfonyl fluoride (PMSF) administration to rats (100 mg/kg, ip in olive oil) as late as 6 or 10 hr after CCl4 (1 ml/kg, ip as a 20% v/v solution in olive oil) can partially prevent the necrogenic response to the hepatotoxin at 24 hr. Here we confirm that observation by electron microscopy and provide further evidence that only in these circumstances were nuclear clumping of chromatin, slight dilatation of the endoplasmic reticulum, myelin figures and lipid droplets in the cytoplasm, large numbers of lysosomes and peroxisomes, glycogen, and slightly swollen mitochondria observable in the protected animals. A very minor part of the late protective effects of PMSF might be due to the effects of this drug on decreasing the intensity of covalent binding of CCl4-reactive metabolites or the intensity of CCl4-induced lipid peroxidation still occurring 6 or 10 hr after CCl4. PMSF administration did not prevent CCl4-induced decreases in cytochrome P450 content or glucose-6-phosphatase activity but partially prevented CCl4-induced calcium accumulation in liver. PMSF treatment increased glutathione and glycogen content in CCl4-poisoned animals, but did not markedly modify protein/phospholipid synthesis or degradation processes. Results suggest that the late protective effects of PMSF administration in CCl4-induced liver necrosis might be due to a favorable modulation of the calcium-calmodulin system similar to that previously described for other drugs.
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PMID:Further studies on the mechanism of the late protective effects of phenylmethylsulfonyl fluoride on carbon tetrachloride-induced liver necrosis. 254 23


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