Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.23.5 (cathepsin D)
 4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activities of a number of peptide-degrading enzymes were compared in homogenates of GH3 cells and rat anterior pituitaries. The enzymes studied were prolyl endopeptidase (EC 3.4.21.26), a soluble metalloendopeptidase, pyroglutamyl peptide hydrolase (EC 3.4.11.8), a multicatalytic protease complex, cathepsin B (EC 3.4.22.1), cathepsin D (EC 3.4.23.5), aminopeptidase (EC 3.4.11.2), and a membrane-bound neutral metalloendopeptidase (EC 3.4.24.11). Specific substrates were used to measure the activities, and active-site-directed inhibitors were used to verify the identities of the enzymes studied. Of the two lysosomal enzymes studied, cathepsin B, the enzyme with the highest activity in both preparations, had 5 times the activity in GH3 cell homogenates as in anterior pituitary homogenates. Cathespin D had a somewhat higher activity in the anterior pituitary homogenates than in the GH3 cell homogenates. Soluble metalloendopeptidase and prolyl endopeptidase, both cytoplasmic enzymes, had about twice the activity in GH3 cell homogenates as in anterior pituitary homogenates. Membrane-bound neutral metalloendopeptidase in the GH3 cell homogenates had 25% of the activity of the anterior pituitary homogenates. Of the two TRH-degrading enzymes, the activity of prolyl endopeptidase in GH3 cell homogenates was about 25 times higher than that of pyroglutamyl peptide hydrolase. Since the secretory function of the pituitary is in part controlled by neuropeptides, the knowledge of the enzyme profiles of the GH3 cells and the anterior pituitary should be of value in studying the metabolism of neuropeptides and peptide hormones in these systems.
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PMID:Peptide-degrading enzymatic activities in GH3 cells and rat anterior pituitary homogenates. 636 4

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

Human beta-secretase candidates, MP78 (h-MP78, EC 3.4.24.15) and cathepsin D (Cat D, EC 3.4.23.5), were evaluated for their ability to enhance amyloid-beta-protein (A beta) secretion when overexpressed in beta APP-containing cells. HEK-293 cells stably co-expressing h-MP78 or Cat D and h-beta APP695 were metabolically labeled with [35S]methionine and A beta secretion was quantified in the conditioned media by immunoprecipitation and ELISA without showing any significant increase in A beta production. Because Cat D is known to have a higher affinity for APP-substrate containing the Swedish familial Alzheimer's disease double mutation (SFAD, K595N and M596L substitutions in beta APP695) than for the wild type substrate [Dreyer et al., Eur. J. Biochem., 224 (1994) 265-271], the effect of Cat D overexpression was tested in a HEK293/beta APPSFAD stable cell line. ELISA analysis of the conditioned media from these cells did also not reveal any increase in A beta generation. In addition, recombinant h-MP78 purified from E. coli cleaved an APP-derived substrate spanning the beta-secretase site (ISEVKMD1AEFRHDS) at multiple sites, but the beta-site cleavage was only a minor one; cleavage occurred predominantly at K-M and E-F bonds. Human liver Cat D also cleaved the same substrate at multiple sites, yet the major cleavage at pH 4.0 occurred at the amyloidogenic D1 site. These findings indicate that h-MP78 does not have the cleavage specificity required for a beta-secretase protease and although Cat D fulfilled the amyloidogenic cleavage specificity, the results of the co-expression experiments make both enzymes less likely candidates as relevant beta-secretases.
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PMID:Expression and characterization of human beta-secretase candidates metalloendopeptidase MP78 and cathepsin D in beta APP-overexpressing cells. 933 17

Huntington's disease (HD) is an inherited and fatal polyglutamine neurodegenerative disorder caused by an expansion of the CAG triplet repeat coding region within the HD gene. Progressive dysfunction and loss of striatal GABAergic medium spiny neurons (MSNs) may account for some of the characteristic symptoms in HD patients. Interestingly, in HD, MSNs expressing neuropeptide Y (NPY) are spared and their numbers is even up-regulated in HD patients. Consistent with this, we report here on increased immuno-linked NPY (IL-NPY) levels in human cerebrospinal fluid (hCSF) from HD patients (Control n = 10; early HD n = 9; mid HD n = 11). As this antibody-based detection of NPY may provide false positive differences as a result of the antibody-based detections of only fragments of NPY, the initial finding was validated by investigating the proteolytic stability of NPY in hCSF using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and selective inhibitors. A comparison between resulting NPY-fragments and detailed epitope analysis verified significant differences in IL-NPY1-36/3-36 and NPY1-30 levels between HD patients and control subjects with no significant differences between early vs mid HD cases. Ex vivo degradomics analysis demonstrated that NPY is initially degraded to NPY1-30 by cathepsin D in both HD patients and control subjects. Yet, NPY1-30 is then further differentially hydrolyzed by thimet oligopeptidase (TOP) in HD patients and by neprilysin (NEP) in control subjects. Furthermore, altered hCSF TOP-inhibitor Dynorphin A1-13 (Dyn-A1-13 ) and TOP-substrate Dyn-A1-8 levels indicate an impaired Dyn-A-TOP network in HD patients. Thus, we conclude that elevated IL-NPY-levels in conjunction with TOP-/NEP-activity/protein as well as Dyn-A1-13 -peptide levels may serve as a potential biomarker in human CSF of HD. Huntington's disease (HD) patients' cerebrospinal fluid (CSF) exhibits higher neuropeptide Y (NPY) levels. Further degradomics studies show that CSF-NPY is initially degraded to NPY1-30 by Cathepsin D. The NPY1-30 fragment is then differentially degraded in HD vs control involving Neprilysin (NEP), Thimet Oligopeptidase (TOP), and TOP-Dynorphin-A network. Together, these findings may help in search for HD biomarkers.
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PMID:Neuropeptide Y (NPY) in cerebrospinal fluid from patients with Huntington's Disease: increased NPY levels and differential degradation of the NPY1-30 fragment. 2701 95