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)

To investigate the role of insulin degrading enzyme (insulysin, EC 3.4.24.56) in insulin signaling, Chinese hamster ovary cells overexpressing the human insulin receptor were genetically engineered to also stably overexpress the rat insulin degrading enzyme. In comparison to the parental cells, these cells expressed 2.7-fold elevated levels of enzyme and insulin degradation was also increased 2-fold. These cells also exhibited a more rapid decrease in receptor tyrosine phosphorylation after removal of insulin. Moreover, low concentrations of insulin were less effective at stimulating proliferation of the cells overexpressing the enzyme. Finally, a fraction of the overexpressed enzyme as well a fraction of the endogenous enzyme could be detected on the plasma membrane surface of these cells. These results support the hypothesis that this enzyme may function in insulin signaling by degrading the insulin molecule.
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PMID:Overexpression of insulin degrading enzyme: cellular localization and effects on insulin signaling. 907 Feb 42

An endoprotease and an aminopeptidase B were isolated from rat testis and characterized. The first one is a metalloendopeptidase of 1161 residues which contains a canonical HXXEHX76E Zn(2+)-binding site and an acidic stretch of 71 amino acids containing 79% of Glu and Asp. It exhibits an in vitro selectivity for peptide bonds at the N-terminus of Arg (R) moieties in dibasic sites and was thus called NRD convertase (Nardilysin: EC 3.4.24.61). It belongs to the pitrilysin family and shows 24 and 34% identity with E. coli protease III (EC 3.4.24.54) and insulysin (EC 3.4.24.55) respectively. The aminopeptidase B component is a 72 kDa metalloexopeptidase which is able to remove Lys and Arg residues from naphtylamide derivatives and from the N-terminus of various peptide substrates. A combination of biochemical and immunochemical studies revealed its ubiquitous character. In the testis, both enzymes are highly expressed at late stages of spermatogenesis and NRD convertase expression is exclusively restricted to the germ cells. The subcellular localization of both enzymes supports the involvement of aminopeptidase B in processing events associated with the secretory pathway but led to new hypothesis on the possible physiological role(s) of NRD convertase.
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PMID:NRD convertase and aminopeptidase B: two processing metallopeptidases with a selectivity for basic residues. 968 93

The malaria parasite Plasmodium falciparum degrades hemoglobin in its acidic food vacuole for use as a major nutrient source. A novel metallopeptidase activity, falcilysin, was purified from food vacuoles and characterized. Falcilysin appears to function downstream of the aspartic proteases plasmepsins I and II and the cysteine protease falcipain in the hemoglobin proteolytic pathway. It is unable to cleave hemoglobin or denatured globin but readily destroys peptide fragments of hemoglobin. Falcilysin cleavage sites along the alpha and beta chains of hemoglobin are polar in character, with charged residues located in the P1 and/or P4' positions. In contrast, plasmepsins I and II and falcipain prefer hydrophobic residues around the scissile bond. The gene encoding falcilysin has been cloned. Its coding sequence exhibits features characteristic of clan ME family M16 metallopeptidases, including an "inverted" HXXEH active site motif. Falcilysin shares primary structural features with M16 family members such as insulysin, mitochondrial processing peptidase, nardilysin, and pitrilysin as well as with data base hypothetical proteins that are potential M16 family members. The characterization of falcilysin increases our understanding of hemoglobin catabolism in P. falciparum and the unusual M16 family of metallopeptidases.
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PMID:Identification and characterization of falcilysin, a metallopeptidase involved in hemoglobin catabolism within the malaria parasite Plasmodium falciparum. 1054 84

Recombinant rat insulysin was shown to cleave the internally quenched fluorogenic peptide 2-aminobenzyl-GGFLRKVGQ-ethylenediamine-2,4-dinitrophenol at the R-K bond, exhibiting a K(m) of 13 microm and a V(max) of 2.6 micromol min(-1) mg(-1). Derivatives of this peptide in which the P(2) leucine or the P(2)' valine were replaced with other residues were used to probe the subsite specificity of the enzyme. Varying the P(2) residue produced a 4-fold range in K(m) and a 7-fold range in k(cat). The nature of the P(2) residue had a significant effect on the site of cleavage. Leucine, isoleucine, valine, and aspartate produced cleavage at the R-K bond. Asparagine produced 36% cleavage at the N-R bond and 64% cleavage at the R-K bond, whereas with alanine or serine the A-R and S-R bonds were the major cleavage sites. With tyrosine, phenylalanine, methionine, or histidine representing the varied residue X, cleavages at F-X, X-R, and R-K were seen, whereas with tryptophan equal cleavage occurred at the F-W and W-R bonds. Variable P(2)' residues produce less of a change in both K(m) and k(cat) and have little influence on the cleavage site. Exceptions are phenylalanine, tyrosine, leucine, and isoleucine, which in addition to producing cleavage at the R-K bond, produce significant cleavage at the L-R bond. Alanine and tyrosine were unique in producing cleavage at the F-L bond. Taken together, these data suggest that insulysin specificity is directed toward the amino side of hydrophobic and basic residues and that the enzyme has an extended substrate binding site.
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PMID:Analysis of the subsite specificity of rat insulysin using fluorogenic peptide substrates. 1104 90

Insulysin (EC. 3.4.22.11) has been implicated in the clearance of beta amyloid peptides through hydrolytic cleavage. To further study the action of insulysin on Abeta peptides recombinant rat insulysin was used. Cleavage of both Abeta(1-40) and Abeta(1-42) by the recombinant enzyme was shown to initially occur at the His(13)-His(14), His(14)-Gln(15), and Phe(19)-Phe(20) bonds. This was followed by a slower cleavage at the Lys(28)-Gly(29), Val(18)-Phe(19), and Phe(20)-Ala(21) positions. None of the products appeared to be further metabolized by insulysin. Using a rat cortical cell system, the action of insulysin on Abeta(1-40) and Abeta(1-42) was shown to eliminate the neurotoxic effects of these peptides. Insulysin was further shown to prevent the deposition of Abeta(1-40) onto a synthetic amyloid. Taken together these results suggest that the use of insulysin to hydrolyze Abeta peptides represents an alternative gene therapeutic approach to the treatment of Alzheimer's disease.
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PMID:Insulysin hydrolyzes amyloid beta peptides to products that are neither neurotoxic nor deposit on amyloid plaques. 1110 81

The intramembranous gamma-secretase cleavage of the beta-amyloid precursor protein (APP) is dependent on biologically active presenilins (PS). Notch also undergoes a similar PS-dependent gamma-secretase-like cleavage, resulting in the liberation of the Notch intracellular domain (NICD), which is critically required for developmental signal transduction. gamma-Secretase processing of APP results in the production of a similar fragment called AICD (APP intracellular domain), which may function in nuclear signaling as well. AICD, like NICD, is rapidly removed. By using a battery of protease inhibitors we demonstrate that AICD, in contrast to NICD, is degraded by a cytoplasmic metalloprotease. In vitro degradation of AICD can be reconstituted with cytoplasmic fractions obtained from neuronal and non-neuronal cells. Taking into account the inhibition profile and the cytoplasmic localization, we identified three candidate enzymes (neurolysin, thimet oligopeptidase, and insulin-degrading enzyme (IDE), also known as insulysin), which all are involved in the degradation of bioactive peptides in the brain. When insulin, a well characterized substrate of IDE, was added to the in vitro degradation assay, removal of AICD was efficiently blocked. Moreover, overexpression of IDE resulted in enhanced degradation of AICD, whereas overexpression of the inactive IDE E111Q mutant did not affect AICD degradation. Finally, immunodepletion of IDE significantly reduced the AICD degrading activity. Therefore our data demonstrate that IDE, which is one of the proteases implicated in the removal of extracellular Abeta, also removes the cytoplasmic product of gamma-secretase cleaved APP.
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PMID:Insulin-degrading enzyme rapidly removes the beta-amyloid precursor protein intracellular domain (AICD). 1180 55

Insulin-degrading enzyme (IDE; insulysin; EC 3.4.24.56) is a 110-kDa neutral metallopeptidase that can degrade a number of peptides including beta-amyloid. The gene encoding IDE is located on chromosome 10 close to a region of linkage for late-onset Alzheimer's disease (LOAD) and thus is a functional and positional candidate for this disorder. We analysed all of the coding exons, untranslated regions and 1000 bp of 5'-flanking sequence of IDE by using denaturing high-performance liquid chromatography and sequencing. We detected eight single nucleotide polymorphisms (SNPs), three in the 5' flanking sequence and five in the coding sequence, of which three were found at lower than 5% frequency. None of them changed the amino acid sequence. We genotyped the five SNPs with allele frequencies of more than 5% in 133 Caucasian LOAD cases and 135 controls collected in the UK and 95 cases and 117 controls collected at the Mayo Clinic, Rochester, USA. Two of the SNPs were analysed in a further independent case-control sample (Washington University, St. Louis: 86 cases, 94 controls). No significant association was found with any individual SNP in any of the samples or with any haplotypes. Analysis of the marker D10S583, which maps 36 kb upstream of IDE, also failed to show association in 134 cases and 111 matched controls from the UK ( P=0.63). Strong linkage disequilibrium was detected between the five SNPs that spanned the whole of the 120-kb genomic region of IDE and one major and a number of minor haplotypes were detected in the populations studied. We conclude that IDE does not make a substantial contribution to the aetiology of LOAD and therefore cannot account for the linkage between LOAD and 10q.
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PMID:Substantial linkage disequilibrium across the insulin-degrading enzyme locus but no association with late-onset Alzheimer's disease. 1181 Feb 77

The steady-state level of amyloid beta-peptide (Abeta) represents a balance between its biosynthesis from the amyloid precursor protein (APP) through the action of the beta- and gamma-secretases and its catabolism by a variety of proteolytic enzymes. Recent attention has focused on members of the neprilysin (NEP) family of zinc metalloproteinases in amyloid metabolism. NEP itself degrades both Abeta(1-40) and Abeta(1-42) in vitro and in vivo, and this metabolism is prevented by NEP inhibitors. Other NEP family members, for example endothelin-converting enzyme, may contribute to amyloid catabolism and may also play a role in neuroprotection. Another metalloproteinase, insulysin (insulin-degrading enzyme) has also been advocated as an amyloid-degrading enzyme and may contribute more generally to metabolism of amyloid-forming peptides. Other candidate enzymes proposed include angiotensin-converting enzyme, some matrix metalloproteinases, plasmin and, indirectly, thimet oligopeptidase (endopeptidase-24.15). This review critically evaluates the evidence relating to proteinases implicated in amyloid catabolism. Therapeutic strategies aimed at promoting A,beta degradation may provide a novel approach to the therapy of Alzheimer's disease.
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PMID:Beta-amyloid catabolism: roles for neprilysin (NEP) and other metallopeptidases? 1206 22

Factors that elevate amyloid-beta (Abeta) peptide levels are associated with an increased risk for Alzheimer's disease. Insulysin has been identified as one of several proteases potentially involved in Abeta degradation based on its hydrolysis of Abeta peptides in vitro. In this study, in vivo levels of brain Abeta40 and Abeta42 peptides were found to be increased significantly (1.6- and 1.4-fold, respectively) in an insulysin-deficient gene-trap mouse model. A 6-fold increase in the level of the gamma-secretase-generated C-terminal fragment of the Abeta precursor protein in the insulysin-deficient mouse also was found. In mice heterozygous for the insulysin gene trap, in which insulysin activity levels were decreased approximately 50%, brain Abeta peptides were increased to levels intermediate between those in wild-type mice and homozygous insulysin gene-trap mice that had no detectable insulysin activity. These findings indicate that there is an inverse correlation between in vivo insulysin activity levels and brain Abeta peptide levels and suggest that modulation of insulysin activity may alter the risk for Alzheimer's disease.
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PMID:Amyloid-beta peptide levels in brain are inversely correlated with insulysin activity levels in vivo. 1273 30

It has been reported previously that ATP inhibits the insulysin reaction (Camberos, M. C., Perez, A. A., Udrisar, D. P., Wanderley, M. I., and Cresto, J. C. (2001) Exp. Biol. Med. 226, 334-341). We report here that with 2-aminobenzoyl-GGFLRKHGQ-ethylenediamine-2,4-dinitrophenyl as substrate, ATP and other nucleotides increase the rate >20-fold in Tris buffer. There is no specificity with respect to the nucleotide; however, ATP is more effective than ADP, which is more effective than AMP. Triphosphate itself was as effective as ATP, indicating it is this moiety that is responsible for activation. The binding of triphosphate was shown to be at a site distinct from the active site, thus acting as a noncompetitive activator. With the physiological substrates insulin and amyloid beta peptide, nucleotides and triphosphate were without effect. However, with small physiological peptides such as bradykinin and dynorphin B-9, ATP and triphosphate increased the rate of hydrolysis approximately 10-fold. Triphosphate and ATP shifted the oligomeric state of the enzyme from primarily dimer-tetramers to a monomer. These data suggest the presence of an allosteric regulatory site on insulysin that may shift its specificity toward small peptide substrates.
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PMID:ATP effects on insulin-degrading enzyme are mediated primarily through its triphosphate moiety. 1549


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