An evolutionarily conserved enzyme degrades transforming growth factor-alpha as well as insulin.

Garcia, J V; Gehm, Barry D.; Rosner, Marsha Rich

doi:10.1083/jcb.109.3.1301

Cited by 51 publications

(31 citation statements)

References 30 publications

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“…Insulysin was shown to be identical to an enzymatic activity referred to as ␥-endorphin-generating enzyme, an enzyme that converts ␤-endorphin to ␥-endorphin (␤Ϫendorphin [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] and ␤Ϫendorphin 1-18 (13). In that study GRF, dynorphin B 1-13, dynorphin A 1-17, and pancreastatin 1-49 were shown also to be substrates.…”

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Analysis of the Subsite Specificity of Rat Insulysin Using Fluorogenic Peptide Substrates

Song

Mukherjee

Juliano

et al. 2001

Journal of Biological Chemistry

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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 M and a V max of 2.6 mol 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.Insulysin (insulin-degrading enzyme; EC 3.4.24.56) was first described as a proteolytic enzyme capable of degrading insulin (1). The enzyme is primarily located in the cytosol and peroxisomes (2), although its presence on the cell membrane (3, 4) and its secretion (5) have recently been reported. Although insulysin has the highest affinity for insulin, the enzyme has been shown to cleave a number of other physiological peptides in vitro including glucagon (6), insulin-like growth factors I and II (7), atrial natriuretic peptide (8), and transforming growth factor-␣ (9). The finding of a variety of substrates for the enzyme, as well as its presence at high levels in insulin-insensitive cells, suggest that insulysin has a variety of physiological functions. Those proposed include processing of insulin for antigen recognition (10), regulation of the multicatalytic proteinase (11), and modulation of steroid receptor action (12).Insulysin was shown to be identical to an enzymatic activity referred to as ␥-endorphin-generating enzyme, an enzyme that converts ␤-endorphin to ␥-endorphin (␤Ϫendorphin 1-17) and ␤Ϫendorphin 1-18 (13). In that study GRF, dynorphin B 1-13, dynorphin A 1-17, and pancreastatin 1-49 were shown also to be substrates. Recent interest in insulysin stems from its ability to degrade the amyloid peptides A␤ 1Ϫ40 and A␤ 1-42 (5, 14 -16) and its possible role as an enzyme involved in the clearance of amyloid peptides in the brain. Decreases in insulysin activity in the brains...

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Analysis of the Subsite Specificity of Rat Insulysin Using Fluorogenic Peptide Substrates

Song

Mukherjee

Juliano

et al. 2001

Journal of Biological Chemistry

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“…In addition to its role in regulating insulin degradation, IDE appears to play an important role in the regulation of growth factor levels and cell differentiation (Becker and Roth, 1995;Authier et al, 1996b). There is, in fact, increasing evidence that IDE binds and degrades transforming growth factor, atrial natriuretic peptide, insulin-like growth factors I and II (Garcia et al, 1989;Müller et al, 1992). IDE is also suggested to regulate extracellular level of amyloid b-protein, accumulation of which is an early and necessary step in the pathogenesis of Alzheimer's disease (Qiu et al, 1998;Kurochkin and Goto, 1994).…”

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Insulin-Degrading Enzyme Exists Inside of Rat Liver Peroxisomes and Degrades Oxidized Proteins.

Morita

Kurochkin

Motojima

et al. 2000

Cell Struct. Funct.

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ABSTRACT. Insulin-degrading enzyme (IDE) was detected by immunoblot analysis in highly purified rat liver peroxisomes. IDE in the peroxisomal fraction was resistant to proteolysis by trypsin and chymotrypsin under conditions where the peroxisomal membranes remained intact. After sonication of the peroxisomal fraction, IDE was recovered in the supernatant fraction. Further, the localization of IDE in the peroxisomes was shown by immunoelectron microscopy. In addition, IDE isolated from peroxisomes degraded insulin as well as oxidized lysozyme as a model substrate for oxidized proteins. These results suggest that IDE exists in an active form in the matrix of rat liver peroxisomes and is involved in elimination of oxidized proteins in peroxisomes.

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“…Although its affinity for insulin is especially high, IDE also has several other substrates in vitro, suggesting a potentially wider role for the enzyme in the clearance of hormones and bioactive peptides. Among these peptide substrates are glucagon, insulin-like growth factors I and II (19), atrial natriuretic factor (20,21), transforming growth factor-␣ (22), and, more recently, growth hormone-releasing factor, ␤-endorphin (23), and the amyloid-␤ protein (A␤) (24). Although IDE's physiological involvement in the degradation of most of these peptides remains hypothetical, the metallopeptidase has recently been shown to participate in the clearance of A␤ in the culture medium of the BV2 microglial cell line (25,26).…”

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Functional Human Insulin-Degrading Enzyme Can Be Expressed in Bacteria

Chesneau

Rosner

2000

Protein Expression and Purification

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An evolutionarily conserved enzyme degrades transforming growth factor-alpha as well as insulin.

Cited by 51 publications

References 30 publications

Analysis of the Subsite Specificity of Rat Insulysin Using Fluorogenic Peptide Substrates

Analysis of the Subsite Specificity of Rat Insulysin Using Fluorogenic Peptide Substrates

Insulin-Degrading Enzyme Exists Inside of Rat Liver Peroxisomes and Degrades Oxidized Proteins.

Functional Human Insulin-Degrading Enzyme Can Be Expressed in Bacteria

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