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:3.4.23.5 (cathepsin D)
4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

No single protease has emerged that possesses all the expected properties for beta-secretase, including brain localization, appropriate peptide cleavage specificity, and the ability to cleave amyloid precursor protein exactly at the amino-terminus of beta-amyloid peptide. We have isolated and purified a brain-derived activity that cleaves the synthetic peptide substrate SEVKMDAEF between methionine and aspartate residues, as required to generate the amino-terminus of beta-amyloid peptide. Its molecular size of 55-60 kDa and inhibitory profile indicate that we have purified the metalloprotease EC 3.4.24.15. We have compared the sequence specificity of EC 3.4.24.15, cathepsin D, and cathepsin G for their ability to cleave the model peptide SEVKMDAEF or related peptides that contain substitutions reported to modulate beta-amyloid peptide production. We have also tested the ability of these enzymes to form carboxyl-terminal fragments from full-length, membrane-embedded amyloid precursor protein substrate or amyloid precursor protein that contains the Swedish KM --> NL mutation. The correct cleavage was tested with an antibody specific for the free amino-terminus of beta-amyloid peptide. Our results exclude EC 3.4.24.15 as a candidate beta-secretase. Although cathepsin G cleaves the model peptide correctly, it displays poor ability to cleave the Swedish KM --> NL peptide and does not generate carboxy-terminal fragments that are immunoreactive with amino-terminal-specific antiserum. Cathepsin D does not cleave the model peptide or show specificity for wild-type amyloid precursor protein; however, it cleaves the Swedish "NL peptide" and "NL precursor" substrates appropriately. Our results suggest that cathepsin D could act as beta-secretase in the Swedish type of familial Alzheimer's disease and demonstrate the importance of using full-length substrate to verify the sequence specificity of candidate proteases.
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PMID:Evaluation of cathepsins D and G and EC 3.4.24.15 as candidate beta-secretase proteases using peptide and amyloid precursor protein substrates. 863 67

The alternatively spliced type III connecting segment (IIICS) of fibronectin (Fn) contains an amino acid sequence, CS-1, which is recognized by the integrin receptor, alpha 4 beta 1. Plasma Fn inhibits alpha 4 beta 1-dependent binding of lymphocytes and monocytes to CS-1 containing Fn derivatives poorly, suggesting limited exposure of the CS-1 sequence in Fn. To test the availability of CS-1 in plasma Fn, an antibody was raised to the synthetic peptide CS-1. The CS-1 sequence was found to be minimally exposed in plasma Fn; and immobilization of Fn, a model of matrix deposition, caused only a modest increase in its exposure. Digestion of Fn with selected proteases, however, induced substantial expression of the CS-1 sequence. The acid protease cathepsin D generated fragments of 31-33.5 kDa from the COOH-terminal heparin-binding domain of Fn which possessed high immunoreactivity with anti-CS-1. Digestion of Fn with cathepsin B also resulted in the exposure of CS-1 sequence in a 140 kDa fragment. Although the digestion of Fn with neutral proteases (neutrophil elastase, cathepsin G, chymotrypsin, trypsin) generated fragments from the COOH-terminal heparin-binding domain of similar molecular weight as with cathepsin D, the exposure of CS-1 did not occur. Exposure of the CS-1 region by the cathepsins was supported by cell adhesion experiments; digestion of Fn with cathepsins D and B transformed inert plasma Fn to an effective inhibitor of adhesion of lymphoblastoid B and T cells (Ramos, Jurkat, Molt-4) to an immobilized CS-1 conjugate. These results suggest that exposure of the CS-1 sequence in plasma Fn by proteolysis with cathepsins D and B, enzymes implicated in several pathological processes, may serve a regulatory function in cell adhesion. The adhesive function of the CS-1 region in intact Fn appears to be suppressed by the native conformation of the molecule.
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PMID:Proteolysis regulates exposure of the IIICS-1 adhesive sequence in plasma fibronectin. 871 84

Regional periprosthetic bone resorption plays an important role of prosthesis loosening. In order to study the possible mechanisms of loosening, we investigated the presence of matrix proteolytic enzymes in the periprosthetic tissue by immunohistochemical technique in 72 patients undergoing revision operation of loosened joint prosthesis, including 22 males and 50 females and aged from 19 to 88 years (mean, 61.7 years). Thirty-nine patients had a loosened hip prosthesis (18 males and 21 females) whereas 33 patients had a loosened knee prosthesis (4 males and 29 females). Tissue specimens collected during revision surgery underwent thin slide sections and H & E staining, and were observed under light microscopy and polarized-light microscopy. The results showed many macrophages, histiocytes, fibroblasts, as well as many phagocytosed metal debris and polyethylene debris in the periprosthetic tissues, suggesting an active bone resorption. Furthermore, we used immunohistochemical techniques to detect the distribution of matrix proteolytic enzymes in periprosthetic tissue, including lysosome enzymes (cathepsin B, cathepsin D and cathepsin G), and matrix metalloproteinase (MMPs, MMP-1, MMP-2, MMP-3). The immunostaining were classified as strong positivity, > 70% positive cells; moderate positivity, 20-70% positive cells; weak/negative, < 20% positive cells. The results showed that cathepsin B, cathepsin D and cathepsin G were found in most fibroblasts and macrophage-like cells, including multinuclear giant cells and epithelioid cells. MMPs were found in most fibroblasts and macrophage-like cells, as well as a scant amount in the extracellular matrix. These enzymes were also found in or around blood vessels, the endothelial cells in the richly vascularized tissue. All negative controls showed no staining. The results of immunoreactive staining ranged from 61.1% to 68.1% of strong to moderate positivity. Since these enzymes were related to the degradation of matrix protein, they may be related to the periprosthetic bone resorption. The further clinical significance needs further investigation.
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PMID:Immunohistochemical analysis of matrix proteolytic enzymes in the periprosthetic tissue in the patients with loosening prostheses. 960 17

Angiotensin II regulates blood pressure and may affect adipogenesis and adipocyte metabolism. Angiotensin II is produced by cleavage of angiotensinogen by renin and angiotensin-converting enzyme in the circulation. In addition, angiotensin II may be produced in various tissues by enzymes of the renin-angiotensin system (RAS) or the nonrenin-angiotensin system (NRAS). We have analyzed the expression of angiotensinogen and enzymes required for its conversion to angiotensin II in human adipose tissue. Northern blot demonstrated angiotensinogen expression in adipose tissue from nine obese subjects. Western blot revealed a distinct band of expected size of the angiotensinogen protein (61 kDa) in isolated adipocytes. RT-PCR, followed by Southern blot, demonstrated renin expression in human adipose tissue. Angiotensin-converting enzyme messenger RNA was detected by RT-PCR, and the identity of the PCR products was verified by restriction enzyme cleavage. Transcripts for cathepsin D and cathepsin G, components of the NRAS, were detected by RT-PCR, verified by restriction enzyme cleavage. We conclude that human adipose tissue expresses angiotensinogen and enzymes of RAS and NRAS. This opens the possibility that angiotensinogen-derived peptides, produced in adipose tissue itself, may affect adipogenesis and play a role in the pathogenesis of obesity.
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PMID:Human adipose tissue expresses angiotensinogen and enzymes required for its conversion to angiotensin II. 981 70

As the amyloidogenic processing of beta-amyloid precursor protein (betaAPP) proceeds under conditions of oxidative stress, the methionine-596 residue at the beta-secretase cleavage point is likely in an oxidized state. In the present work, possible consequences of the oxidation of Met-596 for the generation of the N-terminus of amyloid beta protein were modeled using synthetic peptide substrates, matching 587-606 sequence fragment of betaAPP and containing either intact methionine or methionine sulfoxide. Patterns and rates for the cleavage of these substrates by purified mast cell chymase, cathepsin G, cathepsin D, matrix metalloproteinase-3 and neutrophil elastase, were compared. Only the three first proteases, all previously suggested as candidate beta-secretases, preferentially cleaved the "intact" substrate after Met-596. For chymase and cathepsin G, the specificity of this cleavage increased upon a shift from optimal alkaline pH to acidic pH, which is also more compatible with the plausible intracellular localization of amyloidogenic betaAPP processing. The substitution of methionine sulfoxide for methionine in the substrate slowed down the cleavage rate for all the enzymes tested, by a factor of 6-15. This was associated with shifts of cleavage preferences to points of minor importance for the "intact" peptide, suggesting a specific resistance of the peptide bond after MetSO-596 against proteolysis.
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PMID:Effect of oxidation of beta-amyloid precursor protein on its beta-secretase cleavage. A model study with synthetic peptides and candidate beta-secretases. 983 10

By focusing on the amphiphilic properties of cyclopropenone (e.g. a good electrophile and a precursor for a stable 2pi-aromatic hydroxycyclopropenium cation), a new class of cysteine proteinase inhibitors containing a cyclopropenone moiety was designed. For the purpose of the present research, we needed to devise a new method to introduce a peptide-related moiety as a substituent on the cyclopropenone residue. We investigated the reaction of metalated cyclopropenone acetal derivatives (2, R2 = metal) with N-protected alpha-aminoaldehydes 4 to obtain the adduct 5, and succeeded in the preparation of highly potentiated cysteine proteinase inhibitors 8 after several steps transformations. They showed strong inhibitory activities only to cysteine proteinases such as calpain, papain, cathepsin B, and cathepsin L and not to serine (e.g. thrombin and cathepsin G) and aspartic proteinases (e.g. cathepsin D). Kinetic studies indicated that they are competitive inhibitors, and by the examinations of their inhibitory mechanism it became clear that they are reversible inhibitors.
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PMID:Cyclopropenone-containing cysteine proteinase inhibitors. Synthesis and enzyme inhibitory activities. 1035 36

In the present paper, we demonstrate that alpha1-antichymotrypsin, a serpin with high inhibitory specificity toward cathepsin G, and kallistatin, a human serpin with high specificity toward tissue kallikrein, are digested by cathepsin D. Alpha1-Antichymotrypsin was hydrolyzed essentially in the reactive center loop at L-S, A-L, or L-V bonds; kallistatin was split into small fragments, but we detected the cleavage at F-F and F-S bonds in its reactive center loop in the first 15 min of digestion. In contrast to alpha1-antichymotrypsin, kallistatin is irreversibly inactivated at pH 4.0. Synthetic internally quenched fluorescent peptides containing sequences similar to the reactive center loops of these serpins were hydrolyzed by cathepsin D. The peptides derived from kallistatin were hydrolyzed more efficiently, and particularly relevant was the high susceptibility of the substrates Abz-AIKFFSAQTNRHILRFNRQ-EDDnp (Km = 0.08 microM, kcat = 2.4 s(-1)) and Abz-AIKFFSAQTNRQ-EDDnp (Km = 0.8 microM, kcat = 17.8 s(-1)), which were hydrolyzed at the F-F bond. Therefore, besides the description of a new class of very efficient internally quenched substrates for cathepsin D, we give evidence for the downregulation role of this proteinase on alpha1-antichymotrypsin and kallistatin. The acidification of extracellular milieu by tumor cells can result in activation of cathepsin D; as a consequence, kinins can be released, improving blood supply and leaving more cathepsin G available for the degradation of extracellular matrix.
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PMID:Alpha1-antichymotrypsin and kallistatin hydrolysis by human cathepsin D. 1113 Nov 47

We investigated whether vascular smooth muscle cells (VSMC)-derived from human produce angiotensin (Ang) II upon change from the contractile phenotype to the synthetic phenotype by incubation with fibronectin (FN). Expression of alpha-smooth muscle (SM) actin, apparent in the contractile phenotype, was decreased by FN. Expressions of matrix Gla and osteopontin, apparent in the synthetic phenotype, were increased by FN. Ang II measured by radioimmunoassay (RIA) was significantly increased in human VSMC by FN. Expression of mRNAs for Ang II-generating proteases cathepsin D, cathepsin G, ACE, and chymase was increased by FN. Expressions of cathepsin D and cathepsin G proteins were also increased by FN. Ang I-generating activity, which was inhibited by an aspartyl protease inhibitor pepstatin A, was readily detected in the conditioned medium from human VSMC. Antisense oligodeoxynucleotides (ODNs) that hybridize with cathepsin D and cathepsin G significantly inhibited FN-increased Ang II in conditioned medium and cell extracts. In VSMC conditioned medium, FN-induced elevation of Ang II was significantly inhibited by temocapril but not by chymostatin. Ang II type 1 receptor antagonist CV11974 completely, and antisense cathepsin D and cathepsin G ODNs partially inhibited the FN-stimulated growth of human VSMC. These results indicate that the change of homogeneous cultures of human VSMC from the contractile to the synthetic phenotype sequentially increases expression of proteases cathepsin D, cathepsin G, and ACE, production of Ang II and productions of growth factors, culminating in VSMC proliferation. These findings implicate a new mechanism for the pathogenesis of human vascular proliferative diseases.
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PMID:Human-derived vascular smooth muscle cells produce angiotensin II by changing to the synthetic phenotype. 1281 21

Myeloperoxidase (MPO) is a cationic protein and one of the major constituents of azurophilic granules in neutrophils. Here, we examined whether intracellular transport of MPO and serglycin, a chondroitin sulfate (CS)-bearing proteoglycan, is correlated. First, we examined binding of MPO to CS-Sepharose and measured an ionic interaction, which was disrupted by 200-400 mM NaCl. Next, HL-60 promyelocytes were activated with a phorbol ester, which induced an almost complete rerouting of serglycin from the granular to the secretory pathway, concomitant with a similar effect on MPO transport and secretion. We then used the membrane-permeable cross-linker dithiobis(succininmidylpropionate; DSP) after labeling HL-60 cells with [35S]methionine and [35S]cysteine for 19 h. Immunoprecipitation of MPO revealed its cross-linking to high molecular material having the appearance of a proteoglycan in sodium dodecyl sulfate-polyacrylamide gels. This assumption was confirmed by labeling HL-60 cells with [35S]sulfate for 10 min followed by DSP cross-linking and immunoprecipitation. From three granular enzymes immunoprecipitated, only the cationic MPO was cross-linked to [35S]sulfate-labeled serglycin in appreciable quantities, whereas cathepsin D or beta-N-acetylhexosaminidase was not. Thus, intracellular transport of MPO appears to be linked to that of serglycin. Extracts from high buoyant density organelles from human placenta containing MPO activity were subjected to CS-affinity chromatography. Proteins binding to CS were identified by mass spectrometry as MPO, lactoferrin, cathepsin G, and azurocidin/cationic antimicrobial protein of molecular weight 37 kDa, suggesting that serglycin may be a general transport vehicle for the cationic granular proteins of neutrophils.
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PMID:Targeting myeloperoxidase to azurophilic granules in HL-60 cells. 1296 Feb 44

Stefin B (cystatin B) is an inhibitor of lysosomal cysteine cathepsins and does not inhibit cathepsin D, E (aspartic) or cathepsin G (serine) proteinases. In this study, we have investigated apoptosis triggered by camptothecin, staurosporin (STS), and anti-CD95 monoclonal antibody in the thymocytes from the stefin B-deficient mice and wild-type mice. We have observed increased sensibility to STS-induced apoptosis in the thymocytes of stefin B-deficient mice. Pretreatment of cells with pan-caspase inhibitor z-Val-Ala-Asp(OMe)-fluoromethylketone completely inhibited phosphatidylserine externalization and caspase activation, while treatment with inhibitor of calpains- and papain-like cathepsins (2S,3S)-trans-epoxysuccinyl-leucylamido-3-methyl-butane ethyl ester did not prevent caspase activation nor phosphatidylserine exposure. We conclude that sensitization to apoptosis induced by STS in thymocytes of stefin B-deficient and wild-type mice is not dependent on cathepsin inhibition by stefin B.
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PMID:Sensitization of stefin B-deficient thymocytes towards staurosporin-induced apoptosis is independent of cysteine cathepsins. 1581 33


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