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)

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

Proteoglycans from bovine nasal septa and from the Swarm rat chondrosarcoma were isolated as aggregates (PGC) and as monomers (PGS). Portions of the PGC preparations were degraded with cathepsin D or chondroitinase AC. Chondroitin sulfates were isolated by differential precipitation from alkaline digests of the PGS from bovine nasal septa. The effects of these preparations at concentrations up to 2 mg/ml on the precipitation of tricalcium phosphate in vitro at pH 7.8 in 16 hours at 25 degrees C were ascertained. To this end, the amounts of calcium and phosphate in the precipitates and in the supernates were determined. The PGC preparations were found to be very effective inhibitors; in the presence of 2 mg/ml, precipitate did not form. The PGS preparations were less effective than the PGC preparations; in the presence of 2 mg/ml, about 20% as much calcium phosphate precipitated as in their absence. The chondroitinase AC-degraded preparations at concentrations equivalent to 2 mg/ml of the PGC preparations were approximately as effective as the PGS preparations. On the other hand, the cathepsin D-degraded PGC preparations and the chondroitin sulfate chains were relatively poor inhibitors. Although the viscosity of the solutions may have influenced the rate at which the precipitates settled to the bottom of the tubes, the amounts of the tricalcium phosphate formed were related to the composition and concentration of the proteoglycan preparations.
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PMID:Role of proteoglycans in endochondral ossification: inhibition of calcification. 393 96

A pepsinogen C-like acid protease zymogen was found in Japanese monkey prostate extract and seminal plasma by means of the double immunodiffusion method using rabbit anti-pepsinogen C antiserum, and was purified from the prostate by a combination of ammonium sulfate fractionation, DEAE-Sephacel chromatography, Sephadex G-100 gel filtration, and immunoadsorption to an anti-pepsinogen C column. The zymogen was purified 6,400-fold in a yield of 13.1%. The purified zymogen gave a single band on polyacrylamide gel electrophoresis both in the presence and absence of sodium dodecyl sulfate. The zymogen was converted to the active form by acid treatment at pH 2.8 for 4 h with concurrent reduction of the molecular weight from 41,000 to 36,000. By the double immunodiffusion method, prostate pepsinogen C-like acid protease zymogen, pepsinogen C, lung procathepsin D-II, and their active forms gave a single, fused precipitin line in agar plate with anti-pepsinogen C antiserum, which did not react with cathepsin D and pepsinogen A. Furthermore, the optimal pH of 2.5-3.0, the effect of pepstatin on the activity, and the amino acid compositions were also in good agreement among these three zymogens, showing that they are very similar protease zymogens.
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PMID:Purification of Japanese monkey prostate acid protease zymogen and its identification as a pepsinogen C-like zymogen. 393 48

Rabbit cardiac cathepsin D is initially synthesized as an inactive, apparent molecular weight (Mr) 53,000, pI 6.6 precursor (procathepsin D) that is proteolytically processed during intracellular transport to produce the Mr 48,000 isoforms of active cathepsin D found in cardiac lysosomes. To examine potential proteases responsible for intracellular proteolytic processing, biosynthetically labeled procathepsin D was isolated from rabbit ventricular tissue perfused for 30 min with [35S]methionine. Procathepsin D was then incubated in vitro (40 degrees C, 1-240 min) with active cathepsin D, papain, and cathepsin B, either singly or sequentially, and the reaction products analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and two-dimensional electrophoresis. Incubation of 35S-labeled procathepsin D with active cathepsin D produced a single reaction product (Mr 51,000; pI 6.2). This limited proteolysis occurred at pH 3-5 and was inhibited by pepstatin. Incubation of 35S-labeled procathepsin D with papain or cathepsin B produced a major reaction product (Mr 48,000; pI 6.4) and a minor form (Mr 50,000; pI 6.0). These reactions occurred at pH 4-7 and were inhibited by leupeptin but not pepstatin. Only the Mr 48,000, pI 6.4 products of papain and cathepsin B-mediated proteolysis comigrated with the most basic isoform of active cathepsin D found in cardiac tissue. In addition, the Mr 51,000 intermediate produced by cathepsin D was susceptible to further limited proteolysis by cysteine proteases with resultant production of a Mr 48,000 product. Thus the intracellular proteolytic processing of rabbit cardiac procathepsin D does not result solely from autocatalysis but requires at least one other protease, possibly cathepsin B.
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PMID:Limited proteolysis of rabbit cardiac procathepsin D in a cell-free system. 396 72

Rabbit synovial fibroblasts in monolayer culture secrete a specific collagenase and a neutral endopeptidase into their serum-free culture medium. The rate of secretion of these two enzymes is increased after the ingestion and storage of latex particles within the vacuolar system of the cells. The increased rates of secretion of the neutral enzymes are stable for over 2 wk in the absence of a further phagocytic bout. In constrast there is little change in the extracellular levels of two lysosomal hydrolases, cathepsin D and beta-glucuronidase. The increase in the secretory rates for the two neutral enzymes is related to the number of latex particles ingested by the cells, and increases of up to 12-fold over the nonphagocytosing cultures were observed. A variety of other materials including mycostatin particles and dextran sulfate also induced increases in the secretion of collagenase. These results are discussed in relation to the turnover of connective tissue matrix macromolecules.
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PMID:Stimulation by endocytosis of the secretion of collagenase and neutral proteinase from rabbit synovial fibroblasts. 437 92

The acid-acting proteinase, cathepsin D (EC 3.4.4.23), was purified from extracts of homogenized rabbit lung and beef lung by autolysis at acid pH, acetone and ammonium sulfate fractionation, column chromatography, and isoelectric focusing. Four isoenzymes were obtained from each source. With acid hemoglobin as the substrate, the proteinase from rabbit lung had a pH optimum of 3.0 and that from beef lung had a pH optimum of 3.6. Their activity was not affected by thiol reagents or by Fe(2+), Mn(2+), or Mg(2+). One isoenzyme from rabbit lung was used to immunize a goat, and one from beef lung was used to immunize a rabbit. In immunoelectrophoresis, each resulting antiserum formed a single precipitin line with its homologous enzyme. They cross-reacted with the other three isoenzymes from the same species, but not with any isoenzyme from the other species. At high concentrations, each antiserum completely inhibited the proteolytic activity of its homologous enzyme. The antiserum against rabbit lung cathepsin D also inhibited the proteolytic activity of rabbit peritoneal and pulmonary macrophages. In limited quantities, this antiserum has now been made commercially available and is being used with labeled antibody techniques to identify under a microscope the presence of cathepsin D in macrophages and to study its role in the pathogenesis of tuberculosis.
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PMID:Purification and properties of the cathepsin D types proteinase from beef and rabbit lung and its identification in macrophages. 478 84

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

GAG metabolism was investigated in rats with experimentally induced diabetes. In comparison to control animals, the uptake of 35S-sulfate was diminished in tissues of diabetic animals. Streptozotocin-induced diabetes showed a significant decrease in the content of GAG fractions except that of non-sulfated GAG in liver and kidney which was unchanged as compared to the control group. In rats rendered diabetic by alloxan, non-sulfated GAG increased appreciably in liver and kidney whereas highly sulfated GAG remained unchanged. In the skins of alloxan-diabetic rats both total and sulfated GAG decreased significantly. The activities of liver beta-glucuronidase, beta-N-acetyl glucosaminidase and cathepsin D were significantly increased in rats treated with streptozotocin and alloxan. In streptozotocin-diabetic rats, renal beta-glucuronidase and beta-N-acetyl glucosaminidase activities were reduced while cathepsin D activity was similar to that of controls. The renal beta-N-acetyl glucosaminidase and cathepsin D activities of alloxan-treated rats were not significantly different from normal but their beta-glucuronidase was significantly increased. In the spleen of streptozotocin-diabetic rats all the enzymes were increased except beta-N-acetyl glucosaminidase which remained unaltered. Increased excretion of uronic acid was observed in diabetic groups. These results collectively indicate that both streptozotocin- and alloxan-induced diabetes altered the synthesis and catabolism of GAG.
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PMID:Influence of streptozotocin- and alloxan-induced diabetes on the metabolism of glycosaminoglycans. 624 Jan 83

Renin and cathepsin D were purified by seven-step procedures involving five steps common to both enzymes. These common five steps were extraction of freeze-dried kidney powder in 30% methoxyethanol-water, diethylaminoethyl-cellulose (DEAE-cellulose) batch absorption and elution, pepstatin-aminohexyl-Sepharose chromatography, Sephadex G-100 chromatography, and DEAE-cellulose chromatography. The renin component was purified further by passage through an anti-rat spleen cathepsin D immunoglobulin G-Sepharose (IgG-Sepharose) column followed by carboxymethyl-Sephadex (CM-Sepharose) chromatography which separated two renin components. Cathepsin D activity obtained by the fifth step was purified by passage through an anti-rat kidney renin IgG-Sepharose column followed by DEAE-Sephacel chromatography which separated three cathepsin D components. The homogeneity of renin and cathepsin D preparations was demonstrated by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. The two components of renins showed molecular weights of 42 000 and 36 000 by gel filtration and 38 000 and 36 000 by SDS gel electrophoresis, respectively. They showed isoelectric points of 5.35 and 5.65 by electrofocusing in 5% polyacrylamide gels. Their optimum pHs of enzyme activity were 6.5 as determined by using nephrectomized rat plasma as a substrate. Their specific angiotensin I (Ang I) generation activities were 158 and 146 micrograms of Ang I (microgram of protein)-1 h-1, respectively, which correspond to 1100 and 1020 Goldblatt units (mg of protein)-1 h-1. The three cathepsins showed molecular weights of 41 000, 43 000, and 41 000 by gel filtration and 46 000, 45 000, and 46 000 by SDS gel electrophoresis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Rat kidney renin and cathepsin D: purification and comparison of properties. 636 Feb 7

Val-D-Leu-Pro-Phe-Phe-Val-D-Leu, a specific inhibitor of aspartate proteinases of the pepsin type, was synthesized. Its bonding to activated 6-aminohexanoic acid-Sepharose 4B afforded an affinity support suitable for the purification of human, porcine, and chicken pepsin, human gastricsin, and bovine cathepsin D. These enzymes bind to the support over the pH range 2-5 at 0-1.5 M concentration of NaCl. A buffer at pH greater than or equal to 6, low ionic strength, and containing 20% dioxane can serve as a general desorption agent. The proteinases were isolated from the crude extracts by a single-step procedure in a high degree of purity and in yields exceeding 70%; human pepsin, however, was not separated from human gastricsin. The support does not show any binding capacity for rat plasma renin at pH 7.4 and for some cysteine endopeptidases (cathepsin B, H, and L) at pH 3-5. The cathepsin D preparations isolated by affinity chromatography on the new support and on pepstatin-Sepharose were of the same degree of purity as evidenced by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, N-terminal amino acid sequences, and specific activity.
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PMID:Purification of pepsins and cathepsin D by affinity chromatography on Sepharose 4B with an immobilized synthetic inhibitor. 643 40


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