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)

Insulin degrading enzymes of rat liver cytosol, the so-called insulin and glucagon degrading proteinase (IGP, EC 3.4.23.5), and two forms of the insulin degrading thiol-protein-disulfide oxidoreductase/isomerase (glutathione-insulin transhydrogenase, TPO, EC 1.8.4.2/5.3.4.1) were separated from each other and partially purified on DEAE-Sephadex. The highly purified proteinase was obtained by polyacrylamide gel electrophoresis of the DEAE-Sephadex-purified enzyme fraction and was used to produce monospecific antibodies to the IGP in rabbits. Strong evidence is given that the insulin and glucagon degrading proteinase is an autonomous enzyme existing in addition to the TPO forms in the cytosol of the liver. Combined action of the proteinase and the TPO system on radioiodinated insulin under various conditions in vitro revealed an independent and non-sequential degradation of insulin by these two enzyme systems.
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PMID:The insulin and glucagon degrading proteinase of rat liver. Separation of the proteinase from the thiol-proteindisulfide oxidoreductases. 637 96

Insulin and glucagon degrading proteinase (EC 3.4.23.5) purified from rat liver cytosol was characterized using radioiodinated insulin and glucagon as substrates. Maximum activity for breakdown of both hormones was found at pH 8.1. Thiol blocking reagents as well as indole derivatives inhibit the proteinase, whereas pepstatin, leupeptin, bestatin, elastatinal, antipain, chymostatin and phosphoramidon do not have any effect. Although the Km values and maximal velocities of insulin and glucagon breakdown deviate strongly from each other, the specificity constants (kcat/Km) for both substrates are nearly identical. The insulin and glucagon degrading proteinase, known as a thiol-dependent enzyme, was found to be also a metallo enzyme. Chelating agents, such as EDTA, EGTA, bipyridine and o-phenanthroline show a concentration dependent inhibition. The strongest inhibitor found was o-phenanthroline. Zn++, Co++, Mn++, and to a smaller extent Cd++ and Fe++, are capable of preventing the o-phenanthroline mediated inhibition. Removal of the protein-bound metal(s) results in a nearly total and irreversible loss of enzymatic activity.
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PMID:The insulin and glucagon degrading proteinase of rat liver: a metal-dependent enzyme. 637 97

Cathepsin D (EC 3.4.23.5), the insulin and glucagon degrading proteinase (IGP, EC 3.4.22.-) and the thiol-protein disulfide oxidoreductase (TPO, EC 1.8.4.2, 5.3.4.1) participate in the intracellular protein degradation, the last one also in post-protein-synthetic processing. The distribution of these enzymes was determined in isolated liver parenchymal cells, Kupffer cells and endothelial cells by means of immunochemical methods in order to further characterize these cell types. The cathepsin D content, expressed as microgram enzyme per mg protein, is about 3 fold higher in endothelial cells and about 5 to 24 fold higher in Kupffer cells than in parenchymal cells. This result confirms an earlier report which is based on the activity determination. The TPO concentration is highest in parenchymal cells with half of that concentration in Kupffer cells and one third in endothelial cells. About 0.5% of the total liver protein is represented by this enzyme. The IGP has been found to be totally absent in non-parenchymal cells. It represents, therefore, together with the glucose-6-phosphatase a valuable marker enzyme for parenchymal cells of rat liver.
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PMID:Distribution of thiol-protein disulfide oxidoreductase, insulin-glucagon proteinase and cathepsin D in different cell types of the rat liver. 644 77

Cancer patients have increased insulin resistance in skeletal muscles and probably also in the liver. The insulin production in response to a glucose challenge is decreased. This is associated with decreased glucose uptake in peripheral tissues and increased gluconeogenesis from amino acids, lactate, and glycerol. The correlation between the insulin response to a glucose challenge and the activities of glycolytic and oxidative rate-limiting enzymes in muscle tissue suggests a common denominator for these metabolic alterations. The most prominent feature in alteration of lipid metabolism is a reduction of body fat, probably dependent on increased lipolysis. The released fatty acids are oxidized outside the tumor mass. Species characteristics may be important for the degree of hyperlipidemia. Wasting of the skeletal muscle mass is caused by decreased protein synthesis and probably increased degradation. Anorexia can induce but not entirely explain this altered protein metabolism. Decreased physical activity may be another important factor for the depressed protein synthesis. Total parenteral nutrition (TPN) improves the muscle protein synthesis. The mechanism behind increased fractional degradation of muscle proteins in vitro is not clear, but it may be coupled to increased cathepsin D activity.
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PMID:Metabolism in peripheral tissues in cancer patients. 680 27

Two types of cathepsin D (cathepsins D-I and D-II) were purified from rhesus monkey lung to homogeneity as judged from disc gel electrophoresis. Cathepsin D-I was purified about 2,000-fold with a 5.1% yield while cathepsin D-II was purified about 2,300-fold with a 14.3% yield. Both cathepsins D were rich in the lysosome fraction of the lung, but appeared to be present in part extracellularly. Both showed a molecular weight of about 35,000 on Sephadex G-100 chromatography, and about 41,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cathepsin D-I showed the maximal activity on bovine hemoglobin and albumin at pH 3.4 and 4.0, respectively. It was most stable in the pH range of 5 to 7, but was rather unstable outside this pH range. Cathepsin D-II was quite similar in properties to that from Japanese monkey lung (Moriyama, A. & Takahashi, K. (1978) J. Biochem. 83, 441-451), and was remarkably stable in the pH range of 1-9. Under the conditions used, it retained at least 80% of the original activity when incubated at 37 degrees C for 20 h in this pH range. This stability seems to allow cathepsin D-II to be fairly active even at pH 1.0. Both cathepsins D acted on protein substrates fairly similarly and hydrolyzed hemoglobin most rapidly among the proteins tested. They did not hydrolyze N-acetyl-L-phenylalanyl-3,5-diiodotyrosine. Upon incubation with the oxidized B-chain of insulin, both cathepsins D hydrolyzed the Ala-Leu, Leu-Tyr, Tyr-Leu, Phe-Phe, and Phe-Tyr bonds at both pH 3.0 and 5.0. In addition, cathepsin D-II hydrolyzed the Leu-Val and Tyr-Thr bonds at pH 3.0 and the Val-Asn bond at pH 5.0. Both cathepsins D were inactivated by acid protease-specific inhibitors such as pepstatin, 1,2-epoxy-3-(p-nitrophenoxy)propane, p-bromophenacyl bromide, and diazoacetyl-DL-norleucine methyl ester, although cathepsin D-II was much less susceptible to these reagents except p-bromophenacyl bromide.
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PMID:Cathepsins D from rhesus monkey lung. Purification and characterization. 699 76

When leupeptin, a thiol protease inhibitor of microbial origin, was injected into rats, the activity of fructose-1,6-bisphosphate aldolase (D-fructose-1,6-bisphosphate D-glyceraldehyde-3-phosphate-lyase, EC 4.1.2.13) in the liver decreased to about 60% of that in control rats. However, the concentration of aldolase protein in the liver extracts, measured with a specific antibody obtained with enzyme purified on a phosphocellulose column, remained unchanged. Injection of leupeptin also caused a marked increase in the activities of free lysosomal proteases, such as cathepsin B (EC 3.4.22.1), cathepsin L (EC 3.4.22.-), cathepsin D (EC 3.4.23.5) and lysosomal carboxypeptidase A in the cytosol fraction. A clear inverse relationship between aldolase and cathepsin B activities in the cytosol fraction was demonstrated. The possibility that the less active form of aldolase detected in the livers of leupeptin-treated rats was produced during homogenization was excluded by showing that the aldolase activity was not changed by addition of various protease inhibitors to the homogenization medium., When insulin was coinjected with leupeptin, increase in the activity of free cathepsin L and decrease of activity of aldolase produced by the injection of leupeptin was prevented. These findings indicate that modification of aldolase may be due to the action of a lysosomal protease(s). Enhanced sensitivity of lysosomes to osmotic shock was demonstrated in the livers of leupeptin-treated rats, suggesting that the lysosomal membrane is labilized by administration of leupeptin. Incubation of the purified aldolase with the lysosomal fraction produced the same changes in properties of aldolase as those observed in vivo on injection of leupeptin.
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PMID:Proteolytic modification of rat liver fructose-1,6-bisphosphate aldolase by administration of leupeptin in vivo. 702 Jul 65

Initial cleavage sites of native insulin at a pH of about 3 and stereospecificity were investigated by fungal carboxyl proteinases (EC 3.4.23.6) from ASpergillus sojae, a species of fungi imperfecti, and Pycnoporus coccineus (formerly designated Trametes sanguinea), a wood deteriorating Basidiomycete, respectively. Fungal carboxyl proteinases were used as a model of vertebrate insulin degradation. A. sojae carboxyl proteinase I primarily hydrolyzed two peptide bonds located on the surface of native insulin monomer, the B16-B17 (Tyr-Leu) and B24-B25 (Phe-Phe) bonds, and secondarily the buried bonds, A15-A16 (Gln-Leu), B15-B16 (Leu-Tyr) and B14-B15 (ala-Leu), at pH 3.2 and 30 degree C. The initial cleavage sites of A. sojae carboxyl proteinases I towards native insulin were not identical with the initial cleavage sites towards the oxidized B chain of insulin. P. coccineus carboxyl proteinase Ia selectively hydrolyzed B14-B15 (Ala-Leu), B16-B17 (Tyr-Leu) and B24-B25 (Phe-Phe) bonds in the native insulin at pH 2.7. Based on these findings we suggest that the stereospecificity of the fungal carboxyl proteinases is similar to that of cathepsin D (EC 3.4.23.5), and that the synthesis and degradation of insulin may occur in microorganisms.
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PMID:Initial sites of insulin cleavage and stereospecificity of carboxyl proteinases from Aspergillus sojae and Pycnoporus coccineus. 703 82

Induction of diabetes mellitus in rats following injection of streptozotocin caused reduction in rate of gain of heart weight, of protein and of DNA content in the first two weeks. During the same time interval the overall activity of acid proteinases (cathepsin D), of alkaline proteinases and of proteinase inhibitors was measured in heart muscle homogenates. No statistically significant differences were detected compared with the proteinase activities in control rats. In contrast, total aminopeptidase activity in diabetic hearts was consistently lower than in control hearts. Earlier studies on rat skeletal muscles have shown that induction of diabetes mellitus is followed by a substantial increase of alkaline proteinase as well as aminopeptidase activities. These findings are contrasted by present data obtained with heart muscle of diabetic rats, suggesting that this tissue responds differently to insulin deficiency.
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PMID:Studies on proteolytic activities in heart muscle of diabetic rats. 704 79

In vivo proteolytic modification of liver aldolase on administration of leupeptin, a thiol proteinase inhibitor of microbial origin, is reported. When leupeptin was injected into rats, the activity of aldolase in the liver decreased to 40% of that in control rats. Molecular properties of aldolase isolated from the livers of control rats and leupeptin-treated rats indicated that a decrease of aldolase activity is attributable to hydrolysis of a peptide linkage(s) near the carboxyterminal of the enzyme. Injection of leupeptin also caused marked increase in the activities of free lysosomal proteinases, such as cathepsin A and cathepsin D and moderate increase of cathepsin B and cathepsin L. Increase in free activity of cathepsin A returned to the level of control rats by 12 hr after injection of leupeptin, whereas 36 hr was required for recovery of decreased aldolase activity. When insulin was coinjected with leupeptin, increase in the activity of free cathepsin A and decrease of activity of aldolase produced by the injection of leupeptin was prevented. These findings indicate that modification of aldolase may be due to action of a lysosomal protease(s). Incubation of the purified aldolase with the lysosomal fraction produced the same changes in properties of aldolase as those observed in vivo on injection of leupeptin. The aldolase inactivating proteinase in the lysosomal fraction was inhibited by PMSF and leupeptin and not by pepstatin. Purified cathepsin A (a serine proteinase), cathepsin B and cathepsin L (thiol proteinase) are potent inactivators of aldolase but cathepsin H and cathepsin D are not. Cathepsin A, B and L are involved in inactivation of aldolase in lysosomes. Endogenous thiol proteinase inhibitor which inhibits lysosomal thiol proteinases (cathepsin B, L and H) is found in the cytosol fraction of liver. The level of thiol proteinase inhibitor actually decreased to 60% of that in control rats in leupeptin-treated rats, suggesting that non-thiol proteinase cathepsin A is a major factor in inactivation of aldolase in lysosomes. Not only leupeptin but also other proteinase inhibitors (antipain, E-64-D, chloroquine) caused increase of labilization of the lysosomes and decrease in aldolase activity. Physiological stimuli which are known to induce the labilization of the lysosomal membrane, such as starvation and glucagon, caused slight or no significant increase of activities of free cathepsin A and D and resulted in no apparent change in aldolase activity.
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PMID:Modification of rat liver fructose biphosphate aldolase by lysosomal proteinases. 705 71

The effects of thiols on the breakdown of 125I-labelled insulin, albumin and formaldehyde-treated albumin by highly purified rat liver cathepsins B, D, H and L at pH 4.0 and 5.5 were studied. At both pH values degradation was strongly activated by the thiols cysteamine, cysteine, dithiothreitol, glutathione and 2-mercaptoethanol, and its rate increased with increasing thiol concentration. Preincubation of the protein substrates with 5 mM-glutathione did not affect concentration. Preincubation of the protein substrates with 5 mM-glutathione did not affect the rate of degradation by cathepsin D or L, and determination of free thiol groups after incubation of the proteins in the presence of glutathione but without cathepsin showed that their disulphide bonds were stable under the incubation conditions. Sephadex G-75 chromatography of the acid-soluble products of insulin digestion by cathepsin D or L suggested that thiols can reduce disulphide bonds in proteins after limited proteolysis. The resultant opening-up of the protein structure would lead to further proteolysis, so that the two processes (proteolysis and reduction) may act synergistically. By using the osmotic protection method it was shown that, at a physiological pH, cysteamine, and its oxidized form cystamine, can cross the lysosome membrane and thus may well be the physiological hydrogen donor for the reduction of disulphides in lysosomes. The results are discussed in relation to the lysosomal storage disease cystinosis.
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PMID:Role of thiols in degradation of proteins by cathepsins. 705 70


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