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)

Insulin was enzymatically moniodinated with 127-I or 125-I, and an improved method of purification by anion exchange chromatography was employed. (127-I)Monoiodoinsulin was identified by spectrophotometric analysis and its molar extinction coefficient determined to be 6.31 times 10-3 M-1 cm minus 1. The observed specific activity of carrier-free (125-I)monoidoinsulin was close to the theoretical value (378mCi/mg). The monoiodotyrosyl residue of monoidoinsulin was shown to be solvent-exposed. The ionic properties of the substituted hormone were altered at pH values close to the pK of monoiodotyrosine (8.85), but the pI was unchanged (5.65). (127-I)Monoiodoinsulin formed rhombohedral crystals and co-crystallized with native insulin. Monoidoinsulin was indistinguishable from insulin with respect to binding to two out of three guinea pig anti-insulin sera, to binding to IM9 cultured human lymphocytes, and to binding to isolated rat hepatocyte plasma membranes. The potency of monoidoinsulin was not statistically different from that of insulin in the rat fat cell bioassay and in the mouse convulsion assay. An insulin-degrading enzyme extracted from rat liver degraded monoiodoinsulin less readily than native insulin; monoiodoinsulin was a competitive inhibitor of insulin degradation, and the Km values were 30 nM AND 78 NM for monoidoinsulin and native insulin, respectively. It is concluded that monoidination does not markedly alter the three-dimensional structure of the molecule and that only a few sensitive biological systems are able to distinguish the monoidinated from the native hormone.
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PMID:[127-I]- or carrier-free [125-I]monoiodoinsulin. 23 16

The disappearance rate of intravenously injected insulin was investigated in the serum of 30 women during the third trimester of pregnancy and 6 to 8 weeks post partum, in order to determine whether pregnancy has an influence on insulin kinetics in human subjects. Both women with unimpaired glucose tolerance and those with latent diabetes were included in this study. The disappearance rate of exogenous serum insulin in pregnancy was characterized by a two-compartment model. Multivariate analyses of variance were used to determine whether the estimated parameters of this model during pregnancy differ from those obtained after the puerperium and whether the insulin kinetics are altered when carbohydrate metabolism is disturbed. The kinetics of insulin during pregnancy did not differ from those after pregnancy. Thus, hyperinsulinemia observed in pregnancy cannot be explained by a change in the insulin kinetics. It appears improbable that the insulin-degrading enzyme activities of the placenta participate in degradation of insulin circulating in the maternal blood. A connection between the decline of glucose tolerance during pregnancy and the kinetics of exogenous insulin could not be found.
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PMID:Influence of pregnancy on the kinetics of insulin. 114 34

1. The effects of apomorphine (APO) administration on DA system activity were assessed by measuring dopamine metabolite levels (HVA) in several circumstances. 2. Pretreatment with IMI reduced the effect of APO on HVA levels. 3. Pretreatments with either IDE or DMI did not reduce the effect of APO on HVA levels. 4. Reductions of either NE and 5-HT levels after DSP4 and pCPA restored the effect of APO after IMI pretreatment.
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PMID:Long-term effects of imipramine on striatal dopamine autoreceptor function: involvement of both noradrenergic and serotonergic systems. 138 75

The degradation of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP) by insulin-degrading enzyme (IDE) has been investigated. As revealed by high-performance liquid chromatography, all three peptides are sequentially cleaved at a limited number of sites, the latter of which were identified by mass spectrometric analyses. The studies revealed that ANP is preferred as substrate over BNP and CNP. ANP degradation is rapidly initiated by hydrolysis at the Ser25-Phe26 bond. Three additional cleavage sites were identified in ANP after prolonged incubation with IDE; in contrast, three and two bonds were hydrolyzed in BNP and CNP, respectively. Analysis of the nine cleavage sites shows a preference for basic or hydrophobic amino acid residues on the carboxyl side of a cleaved peptide bond. In contrast to most of the peptide fragments generated by IDE activity, the initial ANP cleavage product, F-R-Y, is rapidly degraded further by cleavage of the R-Y bond. Cross-linking studies with 125I-ANP in the presence of sulfhydryl-modifying agent indicate that IDE activity is inhibited at the level of initial substrate binding whereas metal-ion chelating agents only prevent hydrolysis. On the basis of its structural and enzymatic properties, IDE exhibits striking similarity to a number of recently-described endopeptidases.
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PMID:Rat insulin-degrading enzyme: cleavage pattern of the natriuretic peptide hormones ANP, BNP, and CNP revealed by HPLC and mass spectrometry. 144 54

The degradation of native and 125I-labeled human insulin (HI) was examined in the cytosolic fraction of human, monkey, and rat liver. The purpose of these studies was to provide a species comparison of the interaction of insulin-degrading enzyme (IDE) and protein disulfide isomerase (PDI) in the degradation of HI. Western-blot analysis with monoclonal antibodies indicated the presence of both IDE and PDI in the cytosolic fraction of human and monkey liver. In contrast, rat liver cytosol contained, detectable levels of IDE only. A species comparison of metabolic profiles was performed by fractionating peptide products with reversed-phase high-performance liquid chromatography. After a 60-min incubation, human liver cytosol degraded unlabeled HI into three major products. Two of these peptides coeluted with the products of the incubation of HI with purified rat liver PDI. The three peptides were isolated and determined by NH2-terminal sequence analysis to be intact A chain, B chain, and des(Phe1)-B chain. Human liver cytosol also formed 125I-A chain and 125I-B chain as major products when specifically labeled 125I-HI isomers were used as substrate. Significant proteolytic degradation was observed only when reactions with human liver cytosol were supplemented with Mn2+. In contrast, monkey and rat liver cytosol proteolytically degraded 125I-HI isomers to small peptide fragments. The rat and monkey metabolic profiles were similar to each other and to that observed with Mn(2+)-supplemented human liver cytosol. Proteolysis in monkey and rat was sensitive to inhibition by EDTA.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Mechanisms involved in degradation of human insulin by cytosolic fractions of human, monkey, and rat liver. 160 78

The mechanisms of cellular insulin degradation remain uncertain. Considerable evidence now exists that the primary cellular insulin-degrading activity is a metallothiol proteinase. Two similar degrading activities have been purified and characterized. Insulin protease has been purified from rat skeletal muscle and insulin-degrading enzyme from human red blood cells. Whereas the two degrading activities share a number of similar properties, significant differences have also been reported; and it is not at all established that they are the same enzyme. To examine this, we have compared antigenic and catalytic properties of the two enzymatic activities. Monoclonal antibodies against the red blood cell enzyme adsorb the skeletal muscle enzyme; and on Western blots, the antibodies react with an identical 110-kDa protein. Immunoaffinity-purified enzymes from both red blood cells and skeletal muscle degrade [125I]iodo(B26)insulin to the same products as seen with purified insulin protease and with intact liver and kidney. Chelator-treated muscle and red blood cell enzymes can be reactivated with either Mn2+ or Ca2+. Thus, insulin-degrading enzyme and insulin protease have similar properties. These results support the hypothesis that these activities reside in the same enzyme.
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PMID:Human red blood cell insulin-degrading enzyme and rat skeletal muscle insulin protease share antigenic sites and generate identical products from insulin. 168 96

A metallothiol protease called insulin-degrading enzyme (IDE) seems to be implicated in insulin metabolism to terminate the response of cells to hormone, as well as in other biological functions, including muscle differentiation, regulation of growth factor levels, and antigen processing. In order to obtain highly pure and biologically active IDE, we have developed an immunoaffinity method using a monoclonal antibody to this enzyme (9B12). When the cytosolic fraction of rat liver was first applied to a 9B12-coupled Affi-Gel 10 column, more than 97% of the insulin-degrading activity was absorbed. Among various kinds of buffers successfully eluting the enzyme, only the buffer with a high pH (pH 11) could retain the full biological activity of this enzyme. IDE was further purified via two steps of chromatography using Mono Q anion exchange and Superose 12 molecular sieve columns. The final preparation showed a single band at 110 kDa on reduced sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). In the eluate from the immunoaffinity column, the inhibitory activity associated with the enzyme was also observed. To better recover this endogenous inhibitor, heat-treated cytosolic fraction was fractionated by ammonium sulfate precipitation and applied to the immunoaffinity column on which IDE had been adsorbed. Then, IDE and its inhibitor could be co-eluted with pH 11 as a complex form. After heat treatment of this fraction, the inhibitor was further purified using the same series of chromatography as IDE to more than 20,000-fold; it showed a 14 kDa band on SDS-PAGE. It inhibited both the insulin degradation by IDE in a competitive manner and the cross-linking of 125I-insulin to IDE. Highly purified IDE and the endogenous inhibitor will be useful tools for better understanding the various biological functions of this enzyme.
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PMID:Affinity purification of insulin-degrading enzyme and its endogenous inhibitor from rat liver. 173 Jun 51

The enzymatic and biochemical properties of human insulin-degrading enzyme and Escherichia coli protease III have been compared. Both enzymes were found to degrade insulin in such a way that its receptor binding activity was rapidly lost but its precipitability in trichloracetic acid was only slightly decreased. Both enzymes were also found to be inhibited by chelating agents. The bacterial enzyme, which could be purified in large amounts, was found to contain 0.6 mol of zinc per mol of enzyme but no detectable manganese. The mammalian enzyme but not the bacterial one was inhibited by a sulfhydryl alkylating agent. The two enzymes also differed in substrate specificity. The mammalian enzyme degraded insulin much better than insulin-like growth factor II, whereas the bacterial enzyme degraded them equally. The mammalian enzyme could be labeled by cross-linking to insulin = bombyxin II much greater than insulin-like growth factor I and II much greater than relaxin, while the bacterial enzyme was labeled by insulin-like growth factor II greater than insulin = insulin-like growth factor I much greater than relaxin much greater than bombyxin. Finally, sucrose gradient centrifugation and cross-linking studies both in vitro and in vivo indicated that active human enzyme partially existed as a homo- or heterodimer, whereas the bacterial enzyme was active as a monomer.
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PMID:Comparison of the enzymatic and biochemical properties of human insulin-degrading enzyme and Escherichia coli protease III. 173 42

The insulin-degrading enzyme (IDE) is an evolutionarily conserved enzyme that has been implicated in cellular insulin degradation, but its site of action and importance in regulating insulin degradation have not been clearly established. We addressed this question by examining the effects of overexpressing IDE on insulin degradation in COS cells, using both human IDE (hIDE) and its Drosophila homolog (dIDE). The dIDE, which was recently cloned in our laboratory, has 46% amino acid identity with hIDE, degrades insulin with comparable efficiency, and is readily expressed in mammalian cells. Transient expression of dIDE or hIDE in COS monkey kidney cells led to a 5- to 7-fold increase in the rate of degradation of extracellular insulin, indicating that IDE can regulate cellular insulin degradation. Insulin-degrading activity in the medium was very low and could not account for the difference between transfected and control cells. To further localize the site of IDE action, the fate of insulin after receptor binding was examined. The dIDE-transfected cells displayed increased degradation of prebound insulin compared to control cells. This increase in degradation was observed even when excess unlabeled insulin was added to block reuptake or extracellular degradation. These results indicate that IDE acts at least in part within the cell. The lysosomotropic agents chloroquine and NH4Cl did not affect the increase in insulin degradation produced by transfection with dIDE, indicating that the lysosomal and IDE-mediated pathways of insulin degradation are independent. The results demonstrate that IDE can regulate the degradation of insulin by intact cells via an intracellular pathway.
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PMID:Regulation of insulin degradation: expression of an evolutionarily conserved insulin-degrading enzyme increases degradation via an intracellular pathway. 177 31

A cytosolic protein specifically binding to and degrading atrial natriuretic peptide (ANP) was purified from rat brain homogenate. Based on partial amino acid sequences and enzymatic properties, this protein with an apparent molecular mass of 112 kDa has been identified as the rat insulin-degrading enzyme (IDE). In addition to the known substrates, insulin and transforming-growth-factor alpha IDE binds also with high affinity (apparent Kd 60 nM) to ANP. Competition studies with structural variants of ANP demonstrate that both the C terminus and the disulfide loop of the molecule are essential for high-affinity binding. The data suggest that IDE might be involved in the cellular processing and/or metabolic clearance of ANP.
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PMID:Atrial natriuretic peptide (ANP) is a high-affinity substrate for rat insulin-degrading enzyme. 183 94


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