Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.56 (insulin-degrading enzyme)
 737 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Although insulin-degrading enzyme (IDE) has been implicated in the intracellular degradation of insulin, the cellular localization of this enzyme is still controversial. In the present study, we have examined the cellular localization of IDE in the rat liver by three different techniques using monoclonal antibodies. First, direct immunohistochemical staining of rat liver with one of the monoclonal antibodies revealed that IDE immunoreactivity mainly exists in parenchymal cells, especially in the vicinity of the portal tract and also in the epithelium of the bile duct under light microscopy. In the electron microscopic study, IDE immunoreactivity was found in the cytoplasm near the rough endoplasmic reticulum but not in the plasma membrane, nucleus, or mitochondria. Second, immunoblotting analysis of the subcellular fraction in rat liver showed that the monoclonal antibody specifically reacted with a single polypeptide in the cytosolic fraction, of apparent Mr 110,000, which was consistent with the Mr of IDE. However, a polypeptide band corresponding to IDE could not be observed in the plasma membrane, mitochondrial, or lysosomal fraction. Third, IDE was only detectable in the cytosolic fraction by sandwich radioimmunoassay using two monoclonal antibodies. These results all suggest that IDE is a cytosolic enzyme.
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PMID:Cellular localization of insulin-degrading enzyme in rat liver using monoclonal antibodies specific for this enzyme. 304 64

Although much remains to be learned, our understanding of the mechanisms and processes by which insulin is degraded has advanced considerably over the past few years. The roles of receptor binding and internalization in mediating insulin degradation have been clarified, and the endosomal pathway for intracellular insulin degradation has been established and partially characterized. The importance of IP (IDE) in cellular insulin degradation has been established and the importance of lysosomal degradation questioned. Studies on IP have identified the degradation products resulting from insulin metabolism by this enzyme and shown that the degradation products by IP are identical with those produced by isolated hepatocytes. A major remaining question for future investigation is the potential role of insulin degradation and intracellular processing in insulin action.
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PMID:Insulin degradation: mechanisms, products, and significance. 306 85

The precise mechanism by which insulin is degraded in mammalian cells is not presently known. Several lines of evidence suggest that degradation is initiated by a specific nonlysosomal insulin-degrading enzyme (IDE). The potential importance of this insulin protease is illustrated by the fact that there is an IDE in Drosophila melanogaster Kc cells that shares both physical and kinetic properties with its mammalian counterpart. We now demonstrate that the IDE is present in other Drosophila cell lines and in the embryo, the larvae, the pupae, and adult tissues of the fruit fly. Further, the level of the IDE is developmentally regulated, being barely detectable in the embryo but elevated approximately 5-fold in the larvae and pupae and approximately 10-fold in the adult fly. The IDE levels in the cell lines are particularly high, at least 10-fold greater than in the adult fly. Analysis of Schneider L3 cells indicates that the addition of the Drosophila hormone ecdysone, which induces differentiation of the cells, causes a small but reproducible increase in the level of the IDE and the insulin-degrading activity. These results demonstrate that the IDE is evolutionarily conserved and that its expression is tightly regulated during differentiation of Drosophila. The particular pattern of developmental regulation suggests that the IDE plays a specific and critical role in the later stages of the life cycle of the fly.
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PMID:Developmental regulation of an insulin-degrading enzyme from Drosophila melanogaster. 313 Jun 28

An insulin-degrading enzyme (IDE) from the cytoplasm of Drosophila Kc cells has been purified and characterized. The purified enzyme is a monomer with an s value of 7.2 S, an apparent Km for porcine insulin of 3 microM, and a specific activity of 3.3 nmol of porcine insulin degraded/(min.mg). N-Terminal sequence analysis of the gel-purified enzyme gave a single, serine-rich sequence. The Drosophila IDE shares a number of properties in common with its mammalian counterpart. The enzyme could be specifically affinity-labeled with [125I]insulin, has a molecular weight of 110K, and has a pI of 5.3. Although Drosophila Kc cells grow at room temperature, the optimal enzyme activity assay conditions parallel those of the mammalian IDE: 37 degrees C and a pH range of 7-8. The Drosophila IDE activity, like the mammalian enzymes, is inhibited by bacitracin and sulfhydryl-specific reagents. Similarly, the Drosophila IDE activity is insensitive to glutathione as well as protease inhibitors such as aprotinin and leupeptin. Insulin-like growth factor II, equine insulin, and porcine insulin compete for degradation of [125I]insulin at comparable concentrations (approximately 10(-6) M), whereas insulin-like growth factor I and the individual A and B chains of insulin are less effective. The high degree of evolutionary conservation between the Drosophila and mammalian IDE suggests an important role for this enzyme in the metabolism of insulin and also provides further evidence for the existence of a complete insulin-like system in invertebrate organisms such as Drosophila.
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PMID:Isolation and characterization of an insulin-degrading enzyme from Drosophila melanogaster. 313 25

Histologically the outer layer of the collar enameloid obviously differs from the inner layer, and it has a degree of mineralization nearly as high as the cap enameloid which has the highest. In the stage of matrix formation, the organic matrix of the collar enameloid contains a number of collagen fibers, and odontoblasts display features suggesting that these cells actively synthesized and secreted collagen. A number of cell processes, matrix vesicles and some cell debris which were probably derived from the odontoblasts were observed in the organic matrix of the collar enameloid. We consider that the majority of the organic matrix in collar enameloid originates from the odontoblasts. In the stage of maturation, collagen fibers were not observed in the outer layer of the collar enameloid in demineralized specimens. In the IDE cells during this stage, the complex infoldings of cell membranes developed in the distal portion, and several lysosomal granules and irregular-shaped granules containing many tubular structures, were observed in the distal cytoplasm. In the ODE cells, abundant labyrinthine canals appeared in the cytoplasm, and capillary vessels were found close to the outer surface of the ODE cells. We assume that the higher mineralized outer layer of the collar enameloid is made possible by the absorptive and transport functions of the epithelial cells during the stage of maturation. It is considered that the collar enameloid in this study was initially produced by the odontoblasts and then reconstructed by the epithelial cells, so that the collar enameloid differs from true enamel.
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PMID:The structure and development of the collar enameloid in two teleost fishes, Halichoeres poecilopterus and Pagrus major. 322 8

In the investigation of the intracellular sites of insulin degradation, it might be important whether receptor-bound insulin could be a substrate for insulin-degrading enzyme (IDE). Insulin receptor and IDE were purified from rat liver using a wheat germ agglutinin column and monoclonal anti-IDE antibody affinity column, respectively. [125I]insulin-receptor complex was incubated with various amounts of IDE at 0 degree C in the presence of disuccinimidyl suberate and analyzed by reduced 7.5% SDS-PAGE and autoradiography. With increasing amounts of IDE, the radioactivity of 135 kd band (insulin receptor alpha-subunit) decreased, whereas that of 110 kd band (IDE) appeared then gradually increased, suggesting that IDE could bind to receptor-bound insulin. During incubation of insulin-receptor complex with IDE at 37 degrees C, about half of the [125I]insulin was dissociated from the complex. However, the time course of [125I]insulin degradation in this incubation was essentially identical to that of free [125I]insulin degradation. Cross-linked, non-dissociable receptor-bound [125I]insulin was also degraded by IDE. Rebinding studies to IM-9 cells showed that the receptor binding activity of dissociated [125I]insulin from insulin-receptor complex incubated with IDE was significantly (p less than 0.001) decreased as compared with that without the enzyme. These results, therefore, show that IDE could recognize and degrade receptor-bound insulin, and suggest that IDE may be involved in insulin metabolism during receptor-mediated endocytosis through the degradation of receptor-bound insulin in early neutral vesicles before their internal pH is acidified.
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PMID:Insulin-degrading enzyme is capable of degrading receptor-bound insulin. 327 30

Using conventional techniques of ammonium sulfate fractionation and Sephadex gel column chromatography, insulin-degrading enzyme was partially purified from lysate of human erythrocytes. The enzymatic activity was measured by the trichloroacetic acid precipitation method. Compared to trypsin, the enzyme was highly specific for insulin. The apparent molecular weight of the enzyme was 160,000 Da, the optimum pH was the 7.4 to 7.8 range, and the Km value for insulin for the partially purified enzyme was 162 nM. Bacitracin and N-ethylmaleimide were potent inhibitors, while chloroquine, ethylenediaminetetraacetate, antipain, and soybean trypsin inhibitor failed to inhibit the activity of the enzyme. Like most nucleated cells, human erythrocytes not only have the membranal insulin receptors, but also possess the cytosolic specific insulin-degrading enzyme. Insulin internalization and degradation are shown to be due to the receptor and the enzyme acting in concert as in many nucleated cells. Anucleated erythrocytes have both these entities for possible internalization and degradation of insulin.
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PMID:Characterization of an intracellular insulin-degrading enzyme in human erythrocytes. 329 35

Monoclonal antibodies to a cytosolic insulin-degrading enzyme (IDE) were produced by fusing spleen cells from mouse immunized highly purified human erythrocyte IDE with mouse myeloma cells. Four monoclonal antibodies were identified by their ability to bind to 125I-insulin covalently linked to a cytosolic IDE from human erythrocytes. All four antibodies were found to remove more than 90% of the insulin-degrading activity from erythrocytes extracts, demonstrating that these antibodies were directed against an enzyme which accounts for most of this activity. By immunoprecipitation from metabolically labelled cells and immunoblot procedure, the enzyme from a variety of tissue was shown to be composed of a single polypeptide chain of apparent Mr = 110 kDa. One of these antibodies; 31H7 was coupled to Affi-Gel 10 and used for the purification of this enzyme. Immobilized antigen was eluted at more than 85% efficiency with buffers consisting of either pH2.3, 2.5M MgCl2 or with 6M urea. However, the antigen eluted under 6M urea retained the highest antigenecity (44%) and biological activity (8%) and the yield of the enzyme obtained from this procedure increased up to 17 fold as compared with the conventional method. NaDodSO4/polyacrylamide gel electrophoresis showed a single band of this protein with apparent Mr 110 kDa. These monoclonal antibodies and the purified enzyme will be useful tools for a better understanding of this enzyme, so may lead to the design of specific inhibitors of this enzyme that may be used to treat patients with excessive insulin degradation.
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PMID:[Production of monoclonal antibodies to an insulin degrading enzyme and affinity purification of the enzyme]. 331 32

Clearance of exogenous insulin measured in perfused livers from rats fed ad lib or fasted X 24 or X 48 h was correlated with changes in activity and distribution of the insulin-degrading enzyme glutathione-insulin transhydrogenase measured in microsome fractions, post-perfusion. For comparison with endogenous insulin removal (Endocr. Res. Commun. 7: 231, 1980), a single-pass perfusion mode was used and clearance of insulin at levels (less than or equal to 15 ng/ml) typically observed in perfused rat liver-pancreases during glucose stimulation was studied. Similar to the endogenous data, exogenous insulin removal followed an ogival pattern during fasting. In the fed state, clearance was relatively low, corresponding to a hepatic extraction of approximately 29%. Insulin extraction increased nearly 2-fold after a 24 h fast to approximately 48% (p less than .01), declining to approximately 30% (p less than .025) when fasting was prolonged (X 48 h). At portal insulin concentrations greater than 8 ng/ml (approximately 200 microUnits/ml), clearance tended to decrease in all 3 nutritional states, with apparent saturation of the insulin capturing mechanism being strongest in the 24 h fasted state. In conjunction with these changes in whole organ insulin removal, GSH-insulin transhydrogenase nonlatency, viz., nonlatent activity in intact microsomes relative to total activity in disrupted microsomes, did not change during the first 24 h of fasting; whereas the proportion of nonlatent activity was significantly decreased (p less than .01) after 48 h. Homogenate activity remained essentially constant during the initial fasting period, and declined by approximately 16% (p less than .01) after 48 h of fasting.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Insulin clearance and microsomal glutathione-insulin transhydrogenase in perfused livers of fed and fasted rats. 332 20

Particulate iodipamide ethyl ester, a new hepatolienographic x-ray contrast agent, was intravenously injected into rats. Lung and kidney biopsies taken at various intervals after the injection were examined by light and electron microscopy. IDE particles could be found in the lung capillaries phagocytized by polymorphonuclear neutrophils (PMNs). There were also free particles in the alveolar capillaries in the samples taken 5 min to 4 hours after the injection. No aggregates or emboli were seen. Two days or more after the injection no intra- and extracellular particles were present. The PMNs underwent transient local hydropic degeneration; the lung cells were morphologically intact. In the kidneys, the particles first appeared in both cortical and medullary capillaries. No emboli were observed. The kidney cells did not ingest IDE, but polymorphonuclear neutrophils (PMNs) with ingested IDE were often seen loosely attached to the glomerular capillary walls. In addition, free particles were evident in the capillaries in the samples taken up to 1 hour after injection. All particles in subsequent kidney samples were located in PMNs in the glomeruli. After three or more days the renal tissue was totally devoid of particulate IDE. No morphological evidence of kidney cell injury was observed.
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PMID:Accumulation and elimination of particulate iodipamide ethyl ester in the lungs and kidneys of the rat. A morphological study. 340 91


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