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)

Cryptorchidism of the mature rat testis led to degeneration of the seminiferous tubules and changes in enzyme patterns and activities. Spermatogenic stages 1-4, containing pachytene primary spermatocytes in late meiotic prophase, and stage 5, containing recently formed round spermatids, were damaged by 48 h. Within 96 h stages showed a loss of germinal cells into the lumen and this was almost complete by 192 h. Acid phosphatase showed increased histochemical activity in the basal area of the seminiferous tubule up to 96 h of cryptorchidism, and at 192 h much of the activity was located in large lipidcontaining bodies within the remaining seminiferous epithelium. Total and free biochemical acid phosphatase decreased during cryptorchidism in parallel with cell loss; there were no significant changes in total cathepsin D activity but free enzyme activity was increased throughout the experimental period indicating increased lability of lysosomes in the Sertoli cell. Lactate dehydrogenase activity was mainly tubular but succinate dehydrogenase also showed interstitial activity. Lipoamide dehydrogenase (NADH) was found mainly in the interstitium. During cryptorchidism both lactate and succinate dehydrogenase activity decreased in the tubules parallel to the loss of germinal cells, whereas lipoamide dehydrogenase (NADH) activity increased in both interstitial and tubular areas. It is suggested that the initial lesion in the seminiferous epithelium, produced by cryptorchidism is in the Sertoli cell and that germ cell damage may result from reduced function of the Sertoli cell.
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PMID:The effect of cryptorchidism on the quantitative histology, histochemistry and hydrolytic enzyme activity of the rat testis. 2 15

In populations of cultured arterial endothelial and smooth muscle cells grown under the same conditions, we have measured the total activity per cell of 10 enzymes commonly used as "markers" for subcellular organelles: NADH: ferricyanide reductase, NADH:cytochrome c reductase (rotenone insensitive). NADPH:cytochrome c reductase, alpha-glucosidase, 5'-nucleotidase, alkaline phosphodiesterase I, cytochrome oxidase, monoamine oxidase, cathepsin D, and N-acetyl-beta-glucosaminidase. Significant differences between the cell types were found for 7 of the 10 enzymes tested. The total activity of 5'-nucleotidase in cultured smooth muscle cells was 17 times that of cultured endothelial cells. Comparison of the activities in the two cell types freshly collected and in culture showed that the difference in 5'-nucleotidase in cultured cells is due principally to loss of activity from endothelial cells, suggesting that this activity is regulated differently in the two cell types. In both cell types cathepsin D activity rose during culture.
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PMID:Enzyme activities in endothelial cells and smooth muscle cells from swine aorta. 22 46

Homogenates of HTC cells have been fractionated by differential centrifugation (in four particulate fractions: N, M, L, P, and a supernatant S) or isopycnic banding in linear sucrose gradients. On this basis, the following subcellular organelles may be characterized: (i) Mitochondria, detected by cytochrome oxidase and succinodehydrogenase, are collected in the M and L fractions, and equilibrate, as a narrow band, at a median buoyant density of 1.18 g/cm3. (ii) Lysosomes, detected by the latent hydrolases beta-glycerophosphatase and N-acetyl-beta-glucosaminidase, are largely sedimented in the M and L fractions, and display a broad density distribution pattern with a median value of 1.17 g/cm3. This density is decreased or increased after cultivation of the cells in presence of Triton WR-1339 or Dextran 500, respectively. The behavior of cathepsin D is somewhat at variance with that of the two other hydrolases. (iii) Plasma membrane is tentatively detected by alkaline phosphodiesterase I. Largely recovered in the P fraction, this enzyme equilibrates at a median density close to that of the lysosomal hydrolases; the bulk of cholesterol and about half of the leucyl-2-naphthylamidase are closely associated with alkaline phosphodiesterase I; HTC cells do not contain typical 5'-nucleotidase. (iv) Catalase-bearing particles, of high buoyant density (1.22 g/cm3) are present, but 30-40% of the catalase is also found readily soluble. NADPH- and NADH: cytochrome c reductase, and RNA show more complex distributions. It is suggested that the former enzyme is associated with the endoplasmic reticulum; as in liver, NADH reductase activity is shared between the endoplasmic reticulum and the mitochondria; half of the RNA is associated with free ribosomes of polysomes. True glucose-6-phosphatase could not be detected.
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PMID:Analytical fractionation of cultured hepatoma cells (HTC cells). 56 43

Crystals of the hydrophilic, catalytic domain (30 kDa) of pig liver NADH-cytochrome b5 reductase solubilized by the protease (cathepsin D) have been grown in a solution of polyethylene glycol by the vapor-diffusion procedure. The crystals belong to the orthorhombic system, space group P2(1)2(1)2(1) with unit-cell dimensions of a = 87.1, b = 73.2, and c = 49.0 A. The asymmetric unit contains one molecule of the enzyme. The x-ray diffraction patterns extend to 2.0-A resolution. On the other hand, the intact enzyme (35 kDa) containing the hydrophobic membrane-binding domain solubilized by the detergent (Triton N-101) has been crystallized also from the polyethylene glycol solution. The crystals are needle-shaped and still too small for x-ray diffraction study.
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PMID:Crystallization and preliminary x-ray crystallographic study of NADH-cytochrome b5 reductase from pig liver microsomes. 362 34

Homogenates of cultured rat embryo fibroblasts have been assayed for acid phosphatase, N-acetyl-beta-glucosaminidase, cathepsin D, acid deoxyribonuclease, cytochrome oxidase, NADH cytochrome c reductase, 5'-nucleotidase, inosine diphosphatase, acid pyrophosphatase, neutral pyrophosphatase, esterase, catalase, cholesterol, and RNA. The validity of the assay conditions was checked. Neutral pyrophosphatase is a readily soluble enzyme. Acid hydrolases, except acid pyrophosphatase, are particle-bound enzymes, which exhibit a high degree of structural latency. They are activated and solubilized in a parallel fashion by mechanical treatments and tensio-active agents. Catalase is also particle-bound and latent; activating conditions stronger than those for hydrolases are required to activate the enzyme. Acid pyrophosphatase, 5'-nucleotidase and inosine diphosphatase are firmly particle-bound, but not latent; they are not easily solubilized. In differential and isopycnic centrifugation, the latent hydrolases, cytochrome oxidase and catalase dissociate largely from each other; this suggests the occurrence of lysosomes and peroxisome-like structures besides mitochondria. The distribution patterns of 5'-nucleotidase and cholesterol are largely similar; digitonin influences their equilibrium density to the same extent; these two constituents are thought to be related to the plasma membrane. Inosine diphosphatase and acid pyrophosphatase are also partially associated with the plasma membrane, although some part of these enzymic activities probably belongs to other structures. NADH cytochrome c reductase is associated partly with the endoplasmic reticulum, partly with mitochondria.
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PMID:Analytical fractionation of homogenates from cultured rat embryo fibroblasts. 437 90

The subcellular distribution of rat erythrocyte NADH-cytochrome b5 reductase was determined by radioimmunoassay, using a rabbit antibody against the cathepsin D cleaved water-soluble fragment of rat liver microsomal reductase (I-reductase), which is known to be immunologically similar to the red cell enzyme. Erythrocytes contained approximately 30 ng of reductase/mg of protein, of which 90% were recovered in the hemolysate supernatant and 2.3% in the ghost fraction. After concentration by precipitation with 70% saturated (NH4)2SO4, the NADH-cytochrome c reductase activity of the soluble enzyme could be assayed in the presence of cytochrome b5, and was found to be inhibited by anti 1-reductase antibodies. The sodium dodecyl sulfate-polyacrylamide gel electrophoretic mobilities of erythrocyte membrane-associated and soluble reductase of the liver microsomal enzyme and its cathepsin D cleaved hydrophilic fragment (I-reductase) were examined in crude fractions by blotting followed by specific and highly sensitive immunostaining. The intact microsomal enzyme and the two erythrocyte reductases all had similar mobilities and migrated behind 1-reductase. However, the ghost-associated reductase, which was not attributable to contaminating leukocyte or reticulocyte membranes, was distinguishable from the soluble form by two criteria: (i) a lower dependence on exogenous cytochrome b5 in the NADH-cytochrome c reductase assay; and (ii) a larger apparent Mr upon gel filtration in the presence of Triton X-100, presumably because of detergent binding. Considering these results, possible biogenetic relations between membrane-bound and soluble erythrocyte reductase are discussed.
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PMID:Rat erythrocyte NADH-cytochrome b5 reductase. Quantitation and comparison between the membrane-bound and soluble forms using an antibody against the rat liver enzyme. 714 81

Homogenates of the posterior latissimus dorsi muscle, a phasic muscle, were fractionated by a one-step zonal centrifugation technique into four major organelle populations and cytoplasmic constituents. These were: (1) Plasma membrane fragments with a modal equilibrium density of 1.10 and containing 5'-nucleotidase, alkaline phosphodiesterase, p-nitrophenylphosphatase and acid phosphatase (beta-glycerophosphate was used as the substrate). (2) Sarcoplasmic reticular fragments which could be further subdivided into calcium transport vesicles, with a model equilibrium density of 1.16, that exhibited calcium uptake; K+-ATPase; leucyl-bet-naphthylamidase; acid phosphodiesterase; acid phosphatase (using cytidine monophosphate as the substrate); and sarcoplasmic reticular lysosomes, with a model equilibrium density of 1.18, possessing dipeptidyl-aminopeptidase II, cathepsin D, alpha-glucosidase, N-acetyl-beta-glucosaminidase, and NADH oxidase activity. (3) Mitochondria with a modal equilibrium density of 1.21. (4) Catalase-containing vesicles with a modal equilibrium density of 1.22; and cytoplasmic constituents (modal density of 1.25) with phosphorylase, pyruvate kinase, myosin-ATPase, aldolase, and protein and RNA content. The purity of these organelles was equal to or better than previous efforts, with a 30-fold purification achieved for 5'-nucleotidase and alkaline phosphodiesterase. These results lend support to the hypothesis that the sarcoplasmic reticulum of phasic muscle, in addition to its specialized role in excitation-contraction coupling, represents a multifunctional membrane system, and that, similar to the smooth endoplasmic reticulum of other cells, it includes some membrane-bound lysosomal enzymes and NADH oxidase.
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PMID:Isopycnic-zonal centrifugation of plasma membrane, sarcoplasmic reticular fragments, lysosomes, and cytoplasmic proteins from phasic skeletal muscle. 721 87

The biosynthesis and turnover of rat liver NADH-cytochrome b(5) reductase was studied in in vivo pulse-labeling and long-term, double-labeling experiments. Rats under thiopental anesthesia were injected into the portal vein with [(3)H]L-leucine and sacrificed at various times after the injection. NADH-cytochrome b(5) reductase was extracted from liver cell fractions by cathepsin D-catalyzed cleavage and was then immunoadsorbed onto antireductase-bearing affinity columns in the presence of excess unlabeled rat serum. After elution of the enzyme from the columns with a pH-2.2 buffer, the amount of the reductase protein in the samples was determined by radioimmunoassay, and the radioactivity in reductase was determined on SDS polyacrylamide gel reductase bands. The specific radioactivity of the reductase extracted from the homogenate as well as from rough and smooth microsomal, mitochondrial, and Golgi fractions, estimated at the end of the pulse (10 min after the injection) and at various time points thereafter, remained approximately constant over a 6-h period. These data suggest tha tth eenzyme is independently inserted into the various membranes where it is located. Moreover, the specific radioactivity of the mitochondrial reductase was lower than that of the other fractions, suggesting that it turns over at a slower rate. The lower turnover rate of the mitochondrial enzyme was confirmed by long-term, double-labeling experiments carried out according to the technique of Arias et al. (J. Biol. Chem. 244: 3303-3315.). The relevance of these findings in relation to the understanding of membrane biogenesis and turnover is discussed.
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PMID:Localization and biosynthesis of NADH-cytochrome b5 reductase, an iontegral membrane protein, in rat liver cells. III. Evidence for the independent insertion and turnover the enzyme in various subcellular compartments. 741 81

In this study we present evidence that in human erythrocytes NADH-cytochrome b5 reductase (methemoglobin reductase) is not only soluble but also tightly bound to the membrane. The membrane methemoglobin reductase-like activity is unmasked by Triton X-100 treatment, and represents about half of the total activity in the erythrocytes. Like the amphiphilic microsomal-bound cytochrome b5 reductase, the erythrocyte membrane-bound enzyme is solubilized by cathepsin D. Because this treatment is effective on unsealed ghosts but not on resealed (inside-in) ghosts, it is concluded that the enzyme is strongly bound to the inner face of the membrane. The erythrocyte membrane enzyme is antigenically similar to the soluble enzyme. The two forms of enzyme are specified by the same gene, in that both were found defective in six patients with recessive congenital methemoglobinemia. We suggest that the cytochrome b5 reductase of the erythrocyte membrane is the primary gene product. A posttranslational partial proteolysis probably gives rise to the soluble form of the enzyme, which serves as a methemoglobin reductase.
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PMID:Membrane-bound cytochrome b5 reductase (methemoglobin reductase) in human erythrocytes. Study in normal and methemoglobinemic subjects. 745 47

Reduction of the ascorbate free radical (AFR) at the plasma membrane provides an efficient mechanism to preserve the vitamin in a location where it can recycle alpha-tocopherol and thus prevent lipid peroxidation. Erythrocyte ghost membranes have been shown to oxidize NADH in the presence of the AFR. We report that this activity derives from an AFR reductase because it spares ascorbate from oxidation by ascorbate oxidase, and because ghost membranes decrease steady-state concentrations of the AFR in a protein- and NADH-dependent manner. The AFR reductase has a high apparent affinity for both NADH and the AFR (< 2 microM). When measured in open ghosts, the reductase is comprised of an inner membrane activity (both substrate sites on the cytosolic membrane face) and a trans-membrane activity that mediates extracellular AFR reduction using intracellular NADH. However, the trans-membrane activity constitutes only about 12% of the total measured in ghosts. Ghost AFR reductase activity can also be differentiated from NADH-dependent ferricyanide reductase(s) by its sensitivity to the detergent Triton X-100 and insensitivity to enzymatic digestion with cathepsin D. This NADH-dependent AFR reductase could serve to recycle ascorbic acid at a crucial site on the inner face of the plasma membrane.
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PMID:Recycling of the ascorbate free radical by human erythrocyte membranes. 1142 97


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