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)

In estrogen-receptor-positive human breast cancer cell lines (MCF7, ZR75-1), estrogens specifically increase the secretion into the culture medium of a 52,000 Da (52K) glycoprotein and stimulate cell proliferation. The 52K protein has been purified to homogeneity using monoclonal antibodies and identified as the secreted precursor of a cathepsin D bearing mannose-6-phosphate signals. The secreted precursor 52K protein is mitogenic in vitro in estrogen-deprived MCF7 cells, can be taken up by these cells via mannose-6-phosphate receptors, and can degrade extracellular matrix and proteoglycans following its auto-activation. The protease is also produced constitutively by ER-negative cell lines, and is inducible by tamoxifen in some antiestrogen-resistant variants. The corresponding cDNA has been cloned using N-terminal sequencing of the protein and monoclonal antibodies. Its complete sequencing indicates a strong homology with pro-cathepsin D of normal tissues. Using a cDNA probe, the regulation of 52K cathepsin D mRNA by estrogens and antiestrogens has been studied and chromosome localization determined by in situ hybridization. Clinical studies using both immunohistochemistry and immunoenzymatic assay of breast cancer cytosol have shown that the concentration of total cellular cathepsin D (52K + 48K + 34K) is related to the proliferation of mammary ducts and to the prognosis of breast cancer. Its cytosolic concentration in primary tumors of postmenopausal patients is correlated slightly with lymph node invasion and significantly with shorter disease-free intervals in a 6-year retrospective study with the Danish Breast Cancer Groups and Finsen Institute (S. Thorpe et al.).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Structure, function, regulation and clinical significance of the 52K pro-cathepsin D secreted by breast cancer cells. 314 27

The biochemical properties of renin, extracted from human pituitary specimens obtained at autopsy, were studied using a specific antirenin antibody raised against human kidney renin. The following results were obtained. The molecular weight of pituitary renin was estimated to be about 37,000 daltons by gel filtration through Sephadex G-100. The optimum pH of pituitary renin was between 6.0 approximately 7.0, while that of a renin-like substance which did not react with the antirenin antibody had an acidic pH of 4.0, with a pH comparable to that of the cathepsin D-like enzyme in the pituitary tissue. The presence of two different isoelectric-point species of pituitary renin was revealed by isoelectric focusing, one with a point of pH 4.47 and the other with that of pH 5.77. The Km value of pituitary renin was 37.9 microM for synthetic human renin substrate. Affinity chromatography of the pituitary renin on a Concanavalin-Sepharose column showed that most (87.4%) of the pituitary renin did not contain glycoprotein residues. Treatment with either trypsin or glandular kallikrein increased the renin activity, indicating the presence of an inactive form of renin in the pituitary tissue. From these findings, it is concluded that specific renin exists in human pituitary tissue. It seems likely that the pituitary renin is of local origin rather than contamination of the circulating enzyme.
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PMID:[Biochemical properties of renin in human pituitary tissue]. 389 64

A new aspartic proteinase was isolated from porcine intestine mucosa by affinity chromatography on pepstatin-Sepharose 4B and gel filtration on Sephadex G-100. The enzyme was purified 1600-fold and appeared homogeneous upon polyacrylamide gel electrophoresis. The proteinase has a Mr 60 000 +/- 4000 Da. During sodium dodecyl sulfate polyacrylamide gel electrophoresis the enzyme produced a single protein band (Mr 30 000 +/- 3000 Da). Isoelectric focusing revealed that the enzyme has several multiple forms (pI 6.9, 7.5, 8,0). The enzyme is a glycoprotein containing 5.9% of carbohydrates; the mannose to galactose ratio is 1:3. The amino acid composition of the enzyme was studied. The proteinase splits an oxidized insulin B-chain and synthetic substrates. The pH optimum is 3.2. The enzyme is immunologically identical to porcine spleen cathepsin D.
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PMID:[Proteinases of small intestine enterocytes of swine. Purification and properties of aspartyl proteinase similar to cathepsin D]. 393 2

Multiple biosynthetic forms of glucocerebrosidase were immunoprecipitated after synthesis in vitro using cell-free translation or in vivo using pulse-chase conditions in porcine kidney cells or human fibroblasts. The initial product in vitro was a 52-kDa polypeptide. When canine pancreatic microsomes were present during translation, the nascent polypeptide crossed the microsomal membrane and increased its mass to 60 kDa. Treatment of the 60-kDa polypeptide with endoglycosidase H to remove high mannose carbohydrate yielded a 51-kDa polypeptide. Thus, the membrane-translocated molecule was apparently a high mannose glycoprotein from which a signal peptide had been cleaved, as observed for the lysosomal protease cathepsin D (Erickson, A. H., and Blobel, G. (1979) J. Biol. Chem. 254, 11771-11774). Treatment of pancreatic microsomes or microsomes from porcine kidney cells with protease did not decrease the size of the polypeptide, which shows that this form is not a transmembrane protein bearing a cytoplasmic domain susceptible to digestion. The in vitro product synthesized in the presence of microsomal membranes was indistinguishable from the in vivo product synthesized during pulse-labeling of cultured porcine kidney cells. Following a 2-h chase period, the 60-kDa product was converted to a 59-kDa polypeptide. The major form of glucocerebrosidase detected after a 24-h chase period was a 56-kDa polypeptide, which in turn was converted to a 55-kDa polypeptide by 72 h. The same forms were precipitated from human fibroblasts but the rate of processing was accelerated in this cell type. Limited treatment of the 60-kDa form of glucocerebrosidase with endoglycosidase H suggested that high mannose carbohydrate is added to at least four sites on the polypeptide chain. By 24 h after synthesis, conversion to endoglycosidase H-resistant complex carbohydrate had occurred. Thus, both polypeptide and carbohydrate processing steps are involved in the biosynthesis of glucocerebrosidase.
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PMID:Biosynthesis of the lysosomal enzyme glucocerebrosidase. 393 53

1. Rat kidney lysosomal glycoproteins, prelabelled in the N-acetylneuraminic acid and polypeptide portions with N-acetyl[(3)H]mannosamine and [(14)C]lysine, or with N-acetyl-[(14)C]glucosamine, were incubated under various conditions. Autolytic cleavage of labelled N-acetylneuraminic acid and peptide was maximum at pH5.0. 2. N-Acetylneuraminic acid was released more rapidly than peptide during incubation at 37 degrees or 4 degrees C at pH5. p-Nitrophenyloxamic acid, an inhibitor of bacterial neuraminidase (Edmond et al., 1966), inhibited the cleavage of N-acetylneuraminic acid and peptide, and also inhibited cathepsin D activity. 3. Galactono-, mannono-, and glucono-lactone, inhibitors of the corresponding glycosidases, blocked the autolytic cleavage of N-acetyl[(14)C]glucosamine and protein without inhibiting beta-N-acetylhexosaminidase or cathepsin D activity. These findings suggest that the carbohydrate side chains protect the polypeptide portion of the lysosomal glycoproteins against proteolytic attack by lysosomal cathepsins. 4. In electrofocusing experiments, autolysis was minimized by adding 0.1% p-nitrophenyloxamic acid to the media used for extraction and electrofocusing, and by maintaining an alkaline pH (pH8.8-9) during extraction and dialysis. Arylsulphatase occurred in two forms with pI values of 4.4 and 6.4-6.7, and beta-glucuronidase in two forms with pI values of 4.4 and 6.1. When [(14)C]lysine and N-acetyl[(3)H]mannosamine were given to rats 1.5 and 1 h before killing, (14)C and (3)H were largely restricted to highly acidic glycoprotein species with pI values of 2.1-5.1. 5. When a lysosomal extract was adjusted to pH5 and incubated at 20 degrees C for 16h and then at 37 degrees C for 1 h before electrofocusing, 32 and 58% of the labelled peptide and N-acetylneuraminic acid was cleaved and the pI values of the labelled glycoproteins were markedly increased. About 80% of the acidic form of arylsulphatase and beta-glucuronidase was recovered with the basic form, and the pI of the basic form of both enzymes rose to 7.0. Similar, though less marked changes, were observed when a lysosomal extract was kept at pH5 for 2h at 4 degrees C before electrofocusing. 6. When an acidic lysosomal fraction (pI4.2-4.6) was incubated at pH5 for 2.5h and refocused, 80% of the arylsulphatase now occurred in two forms with pI values of 5 and 6.4. When a basic lysosomal fraction (pI5.8-6.4) was similarly incubated, the pI of arylsulphatase increased from 6.4 to 7.2. The relative increase in pI of arylsulphatases was accompanied by a proportional loss of N-acetylneuraminic acid from the glycoprotein associated with these forms. 7. These experiments show that lysosomal glycoproteins and two representative hydrolases, when exposed to a mildly acidic pH, readily undergo autolytic degradation and their pI values increase. These observations may have a bearing on the origin of the molecular heterogeneity of the lysosomal enzymes.
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PMID:Autolysis of glycoproteins in rat kidney lysosomes in vitro. Effects on the isoelectric focusing behaviour of glycoproteins, arylsulphatase and beta-glucuronidase. 445 20

Cathepsin H was purified from human liver by a method involving autolysis and acetone fractionation, and chromatography on DEAE-cellulose, Ultrogel AcA 54, hydroxyapatite and concanavalin A-Sepharose. The procedure allowed for the simultaneous isolation of cathepsin B and cathepsin D. Cathepsin H was shown to consist of a single polypeptide chain of 28 000 mol.wt., and affinity for concanavalin A-Sepharose indicated that it was a glycoprotein. The enzyme existed in multiple isoelectric forms, the two major forms having pI values of 6.0 and 6.4; it hydrolysed azocasein (pH optimum 5.5), benzoylarginine 2-naphthylamide (Ba-Arg-NNap), leucyl 2-naphthylamide (Arg-NNap), (pH optimum 6.8). Arg-NNap and Arg-NMec, unlike Bz-Arg-NNap-, were not hydrolysed by human cathepsin B. Cathepsin H was similar to cathepsin B in being irreversibly inactivated by exposure to alkaline pH. Sensitivity to chemical inhibitors by 1 microM-leupeptin, which gave essentially complete inhibition of the other lysosomal cysteine proteinases, cathepsins B and L.
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PMID:Human cathepsin H. 616 52

Candida albicans was able to produce a keratinolytic proteinase (KPase) when cultivated in a medium containing human stratum corneum as a nitrogen source. The KPase was purified to 108.5-fold by ion-exchange chromatography and gel filtration. The molecular weight of the enzyme was estimated to be 42,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration through Sephacryl S-200, while the isoelectric point was determined to be at pH 4.5. The enzyme had an optimum pH of 4.0 and was "inactive" below pH 2.5 and above pH 6.0. The activity of KPase after preincubation at various temperatures was stable up to 50 degrees C. The keratinolytic activity was not affected by the addition of nonionic detergents and divalent cations. The enzyme was a glycoprotein and contained a high content of aspartic acid residues (172/1000). Pepstatin and chymostatin inhibited the activity in a dose-dependent manner; however, neither the other group specific inhibitors tested nor the pepsin specific inhibitors, DAN or EPNP, showed any effect on the enzyme. From these inhibitory profiles, this enzyme was determined to be a carboxyl proteinase such as cathepsin D. Among the various substrates for proteolytic enzymes, KPase digested human stratum corneum as much as albumin and hemoglobin. In the three fractions (water soluble, keratin filamentous, and membranous) prepared from human stratum corneum, the keratin filamentous fraction was more susceptible to degradation by KPase than the other two fractions were. KPase also digested much less human fingernail (13%) than human stratum corneum, but did not show any signs of there being any digestion of human scalp hair. These studies suggest that KPase from C. albicans may play an important role in superficial infection by affecting the human stratum corneum of the skin and nail.
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PMID:Isolation and characterization of proteinase from Candida albicans: substrate specificity. 620 88

Cathepsin D was isolated from human brain. A consecutive use of affinity chromatography on hemoglobin-sepharose 4B and column chromatography on hydroxylapatite resulted in a homogeneous enzyme (as was demonstrated by SDS polyacrylamide gel electrophoresis) with a molecular weight of about 48,000, 2800-fold purification and 3.4% yield. Incubation of serum proteins in the presence of purified cathepsin D resulted in a gradual decrease of immunoreactive forms of albumin, orosomucoid, transferrin, and other alpha 1, alpha 2 and beta-globulins. The degradation was revealed by crossed immunoelectrophoresis. Crossed affinity immunoelectrophoresis in the presence of ConA showed specific degradation of serum glycoproteins. Rocket immunoelectrophoresis with monospecific antisera raised against human adult brain glycoprotein D2 revealed a rapid and linear degradation of detergent-solubilized and partially purified human membrane glycoprotein D2 by purified cathepsin D. Incubation of glycoprotein D2 in the presence of cathepsin D (30 min, 37 degrees C) resulted in degradation of 95% of specific protein. An exposure of human brain membrane fragments to cathepsin D resulted in linear degradation of membrane-bound glycoprotein followed by an appearance of a soluble immunoreactive form of protein D2.
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PMID:[Immunochemical study of the degradation of circulating glycoproteins and the neurospecific membrane glycoprotein D2 by cathepsin D of the human brain]. 647 83

We have analyzed a soluble form of the glycoprotein (G) obtained from vesicular stomatitis virus (VSV) by treatment of intact virions with cathepsin D. This form lacks the carboxy-terminal and membrane-spanning domains and thus is analogous to the previously described secreted form of G, Gs. The molecular weight of the cathepsin D produced G, G(Cath D), measured by sedimentation equilibrium in the analytical ultracentrifuge is 57 600, indicating that it is a monomer. Intact G protein extracted from virions by octyl beta-D-glucoside also is monomeric, based on sedimentation equilibrium analysis. These results suggest that G may be monomeric in virions. The Stokes radii (Rs) of the two forms of G were obtained from their migration in nondenaturing polyacrylamide gradient gels. The Rs of G(Cath D) in the absence of nonionic detergent was 37 A; in the presence of nonionic detergent, it increased to 55 A. The Rs of detergent-extracted intact G was 63 A in nonionic detergent. From the molecular weight and Rs of G(Cath D), we calculated a sedimentation coefficient of 3.8 S; the value determined by centrifugation in a sucrose gradient was 3.7 S. Viruses such as VSV fuse with cell membranes at low pH [White, J., Matlin, K., & Helenius, A. (1981) J. Cell Biol. 89, 674-679]. We have used the fluorescent probe cis,trans,trans,cis-9,11,13,15-parinaric acid (cis-PnA) to detect a reversible conformational change in G(Cath D) when the protein was exposed to an acidic environment close to pH 5. cis-PnA binds to hydrophobic regions of protein, causing a quenching of the intrinsic tryptophan fluorescence and an increase in the fluorescence of the probe.
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PMID:Physical properties of a soluble form of the glycoprotein of vesicular stomatitis virus at neutral and acidic pH. 666 13

The ultrastructural cytochemical reactivity, renin activity, and cathepsin D activity of atria and ventricle of the bullfrog have been assessed. The specific granules (A, B, and D) were found to be argentaphobic when ultrathin sections of Araldite-embedded atria and ventricle were stained according to the periodic acid-thiocarbohydrazide-silver proteinate technique of Thiery. The entire core of the specific granules was moderated positive after ultrathin sections of glutaraldehyde-fixed, glycol methacrylate embedded atria and ventricle were stained by phosphotungstic acid at a low pH. A similar reaction was shown by the cell coat, intercalated discs, residual bodies (C granules), and Z discs as well as by a very small portion of the Golgi complex. Incubation of ultrathin sections of atria and ventricule fixed only in glutaraldehyde and embedded in glycol methacrylate with either pronase or trypsin resulted in selective digestion of specific granules and Z discs and, to a much lesser degree, of the cell coat. As cathepsin D activity and renin activity were present in both atria and ventricle, the generation of angiotensin I by these cardiocytes might have been due to either enzyme. Nevertheless, because of the glycoprotein nature of specific granules and of the endocrinelike ultrastructure of atrial and ventricular cardiocytes in the frog, the present results raise the possibility that specific granules may contain renin.
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PMID:Ultrastructural cytochemistry of atrial and ventricular cardiocytes of the bullfrog (Rana catesbeiana). Relationship of specific granules with reninlike activity of the myocardium. 701 73


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