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)

Chinese hamster ovary cells transfected with human lysozyme cDNA encoding Asn instead of Gly22 synthesize a mutant lysozyme, [Asn22]lysozyme, with about 60% of the molecules bearing carbohydrate. This carbohydrate is predominantly of the complex type and contains a varied number of lactosamine repeats. In this study we show that the glycosylation of [Asn22] lysozyme fused to human cathepsin D is altered relative to [Asn22]lysozyme alone. The fusion protein is synthesized as a 66-kDa precursor that is cleaved to enzymatically active and antigenically positive cathepsin D and lysozyme. As compared with [Asn22]lysozyme the lysozyme moiety of the fusion protein shows an increased N-glycosylation and a decreased synthesis of lactosamine repeats. Cleavage of the precursor with cathepsin L has revealed that the lysozyme portion of the secreted fusion protein bears a complex type carbohydrate. The intracellularly released lysozyme portion of the fusion protein contains trimmed oligosaccharides. In the presence of NH4Cl the lysosomal targeting of the fusion protein is inhibited. The secreted protein is then enriched in molecules bearing phosphorylated high mannose oligosaccharides in their lysozyme moiety. Our results indicate that carbohydrate processing in [Asn22]lysozyme, including the synthesis of mannose 6-phosphate residues and of lactosamine repeats, is altered by the attached cathepsin D. The phosphorylation of the carbohydrate on the lysozyme portion results in a very efficient lysosomal targeting of the concerned fusion protein molecules.
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PMID:Synthesis of phosphorylated oligosaccharides in lysozyme is enhanced by fusion to cathepsin D. 836 10

Mannose 6-phosphate (Man-6-P) is a posttranslational carbohydrate modification typical of newly synthesized acid hydrolases that signals targeting from the Golgi apparatus to the lysosome via Man-6-P receptors (MPRs). Using iodinated cation independent MPR as a probe in a Western blot assay, we surveyed levels of Man-6-P glycoproteins in a number of different rat tissues. Considerable variation was observed with respect to total amounts and types of Man-6-P glycoproteins in the different tissues. Brain contained 2-8-fold more Man-6-P glycoproteins than other tissues, with relative abundance being brain >> testis approximately heart > lung approximately kidney approximately ovary approximately spleen > skeletal muscle approximately liver approximately serum. Analysis of 16 different lysosomal enzyme activities revealed that brain contains lower activities than other tissues which suggested that decreased removal of Man-6-P results in increased levels of Man-6-P glycoproteins. This was directly demonstrated by comparing activities of phosphorylated lysosomal enzymes, purified by immobilized MPR affinity chromatography, with total activities. The phosphorylated forms accounted for a considerable proportion of the MPR-targeted activities measured in brain (on average, 36.2%) but very little in lung, kidney, and liver (on average, 5.5, 2.3, and 0. 7%, respectively). Man-6-P glycoproteins were also isolated from rat brain by MPR affinity chromatography on a preparative scale. Of the 18 bands resolvable by SDS-polyacrylamide gel electrophoresis, seven bands were NH2-terminally sequenced and identified as the known lysosomal enzymes cathepsin L, cathepsin A, cathepsin D, alpha-galactosidase A, arylsulfatase A, and alpha-iduronidase. One of the major Man-6-P glycoproteins was identified as palmitoyl protein thioesterase, which was not previously thought to be lysosomal. This finding raises important questions about the cellular location and function of palmitoyl protein thioesterase, mutations in which result in the neurodegenerative disorder, infantile neuronal ceroid lipofuscinosis.
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PMID:Rat brain contains high levels of mannose-6-phosphorylated glycoproteins including lysosomal enzymes and palmitoyl-protein thioesterase, an enzyme implicated in infantile neuronal lipofuscinosis. 870 98

Urokinase plasminogen activator (uPA) is a multifunctional protein involved in both extracellular proteolysis and signal transduction. uPA usually mediates its actions while attached to a membrane-bound receptor, termed uPAR. In this study, uPA and its receptor were measured at both protein and mRNA levels in breast cancer. At both levels, concentrations of uPA were significantly correlated with those for uPAR. uPA levels also correlated significantly with cathepsin B and cathepsin D but not with cathepsin L, MMP-8 or MMP-9 levels. Irrespective of the cut-off point used (e.g., median, tertile or quartile values), uPA was a significant prognostic marker for breast cancer.
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PMID:Urokinase plasminogen activator as a predictor of aggressive disease in breast cancer. 879 99

To estimate the prognostic value of cathepsins B, H, L, D and stefins A and B in head and neck carcinoma, their concentrations in cytosols of primary tumours and adjacent normal tissue were measured (cathepsins B, D stefins A, B in 45, cathepsin L in 24 and cathepsin H in 21 patients). Median concentrations of cathepsins B, L, and D were significantly higher in tumour than in the adjacent normal tissue (B and D: p < 0.0001; L: p = 0.004); cathepsin H concentration was higher in normal tissue (p = 0.001). Concentrations of either stefin did not differ significantly between normal and tumour tissue. Concentrations of cathepsins B, H, L, and D were higher in laryngeal than in non-laryngeal normal and tumour tissues. The difference was statistically significant for cathepsin B in tumour tissue (p = 0.045), and marginally significant in normal tissue (p = 0.07). Early tumours had lower concentrations of stefins A and B than locally advanced tumours (stefin A: p = 0.04; stefin B: p = 0.07). Disease-free and disease-specific survival rates were better in patients with concentrations of cathepsin L in tumour tissue below or equal to the cut-off values (p = 0.035; p = 0.05), whereas for cathepsin B the difference was established only for disease-free survival (p = 0.07). The opposite was true for stefin A (p = 0.0002; p = 0.002) and stefin B (p = 0.009; p = 0.003), and in disease-free survival also for cathepsin H (p = 0.055). The concentration of cathepsin D did not correlate with survival. Our data indicate that cathepsins B, H, L and stefins A and B might have prognostic value in head and neck carcinoma.
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PMID:Prognostic value of cathepsins B, H, L, D and their endogenous inhibitors stefins A and B in head and neck carcinoma. 883 84

Proteinase activity is increased in psoriatic epidermis. To elucidate the involvement of enzymes in psoriatic epidermis, the expression of cathepsins, L, B and D was investigated by Western blotting and immunohistological studies. Normal epidermis contained abundant inactive precursors (39 kDa) of cathepsins L and B and an inactive intermediate form (45 kDa) of cathepsin D. Cathepsin L in psoriasis was processed to a variable extent from the precursor to a single-chain form (30 kDa) and a mixture of single- and heavy-chain (25 kDa) forms of the active mature enzyme, corresponding to the immunohistological staining patterns 'diffuse dense', 'small granular', and unevenly distributed 'condensed granular'. Cathepsin B showed a mixture of precursor form (39 kDa) and single-chain (30 kDa) forms and was expressed as a 'diffuse dense' staining pattern in the mid-spinous layer and as a 'condensed' pattern in the upper spinous and granular layers. Cathepsin D was processed to the heavy-chain (31 kDa) form of activated mature enzyme with small granular staining and a mixture of heavy-chain and degraded protein (28 kDa) with larger and more condensed granular staining. The distribution patterns of 'small granular' cathepsin L, and of cathepsins B and D expression in suprabasal keratinocytes were very similar to that of involucrin. After complete clinical resolution of psoriasis by 8-methoxypsoralen plus UVA treatment, the expression of the three cathepsins was normalized. These results suggest that cathepsins L, B and D in forms activated to a variable extent may be involved in the pathology of psoriasis.
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PMID:Processing of cathepsins L, B and D in psoriatic epidermis. 904 42

We have determined the primary cleavage sites in the bone Gla protein (BGP; osteocalcin) for several of the proteases that could act on the protein during bone resorption and turnover, cathepsins B, D, L, H, and S. The time course of BGP digestion by each cathepsin was first determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. We then incubated human and bovine BGP with each cathepsin for a sufficient time to reduce the level of intact protein by at least 20-fold, isolated the major cleavage peptides, and identified each by N-terminal sequence analysis and by amino acid analysis. Our results show that BGP has relatively few cathepsin-sensitive sites and that these sites are located at the N and C terminus of the 49-residue protein. Cathepsins B, L, H, and S readily cleave BGP at the G7-A8 bond; cathepsin L also cleaves at R43-R44; cathepsin B also cleaves at R44-F45; and cathepsin D cleaves only at A41-Y42. The immunoreactivity of the major peptides generated by cathepsin cleavage was evaluated using the original radioimmunoassay developed for the detection of BGP in human serum. The BGP 8-49 fragment cross-reacts identically with native BGP, while the 8-43 and the 1-44 fragments require 20- to 40-fold higher concentrations to achieve the same level of displacement as the native protein. The 1-41 and 8-41 fragments are unable to significantly displace the labeled native BGP tracer at any concentration tested. These results demonstrate the utility of peptides generated by cathepsin digestion in the mapping of the antigenic epitopes recognized by a given BGP immunoassay.
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PMID:Identification of peptide fragments generated by digestion of bovine and human osteocalcin with the lysosomal proteinases cathepsin B, D, L, H, and S. 907 88

In the seminiferous epithelium, germ cell development behind the blood-testis barrier involves continual degradation and renewal of inter-testicular cell junctions. This allows: (i) the translocation of developing germ cells from the basal lamina to the adluminal compartment during spermatogenesis, and (ii) the eventual release of mature spermatids into the tubular lumen during spermiation. Throughout spermatogenesis, cellular debris must also be removed from the epithelium Thus, it is conceivable that proteases, protease inhibitors, and cell junctional components are involved in these events. The present study sought to examine whether testicular cells can express multiple cathepsin mRNAs given that these proteases are involved in the degradation and processing of proteins as well as in tissue regeneration. By using total RNA isolated from primary cultures of Sertoli, Leydig, and germ cells for reverse-transcription and polymerase chain reaction (RT-PCR), the mRNAs of cathepsin B, C, D, H, L, and S were shown to be expressed by Sertoli and Leydig cells, whereas germ cells isolated from adult rats expressed all of the above cathepsin mRNAs except cathepsin D. Throughout postnatal development and maturation, the testicular steady-state mRNA levels of cathepsin B, C, D, L, and S remain relatively unchanged with the exception of cathepsin H whose mRNA level increased during maturation and peaked at 45-60 days of age. Using lonidamine, an anti-spermatogenic drug which is known to induce premature release of germ cells without affecting Leydig cell function by disrupting the inter-Sertoli-germ cell junctions, we have examined the differential expression of these cathepsin mRNAs in the testis at the time of extensive tissue restructuring. It was noted that the expression of cathepsin L and S in the testis increased significantly concomitant with the disappearance of elongate spermatids whereas the expression of cathepsin B, C, D, and H increased significantly when most of the round spermatids and spermatocytes were depleted. These results illustrate the intricate inter-relationship between these proteases in the testis during maturation and tissue restructuring.
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PMID:Differential expression of multiple cathepsin mRNAs in the rat testis during maturation and following lonidamine induced tissue restructuring. 923 20

The cytotoxicity of oxidized LDL (oxLDL) to several types of artery wall cells might contribute to atherosclerosis by causing cell death, presumably by both apoptosis and necrosis. After its uptake into macrophage lysosomes by receptor-mediated endocytosis, oxLDL is poorly degraded, resulting in ceroid-containing foam cells. We studied the influence ofoxLDL on lysosomal enzyme activity and, in particular, on lysosomal membrane stability and the modulation of these cellular characteristics by HDL and vitamin E (vit-E). Unexposed cells and cells exposed to acetylated LDL (AcLDL) were used as controls. The lysosomal marker enzymes cathepsin L and N-acetyl-beta-glucosaminidase (NAbetaGase) were biochemically assayed in J-774 cells after fractionation. Lysosomal integrity in living cells was assayed by the acridine orange (AO) relocation test. Cathepsin D was immunocytochemically demonstrated in J-774 cells and human monocyte-derived macrophages. We found that the total activities of NAbetaGase and cathepsin L were significantly decreased, whereas their relative cytosolic activities were enhanced, after oxLDL exposure. Labilization of the lysosomal membranes was further proven by decreased lysosomal AO uptake and relocation to the cytosol of cathepsin D, as estimated by light and electron microscopic immunocytochemistry. HDL and vit-E diminished the cytotoxicity of oxLDL by decreasing the lysosomal damage. The results indicate that endocytosed oxLDL not only partially inactivates lysosomal enzymes but also destabilizes the acidic vacuolar compartment, causing relocation of lysosomal enzymes to the cytosol. Exposure to AcLDL resulted in its uptake with enlargement of the lysosomal apparatus, but the stability of the lysosomal membranes was not changed.
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PMID:Uptake of oxidized LDL by macrophages results in partial lysosomal enzyme inactivation and relocation. 948 81

Incubation of cultured hippocampal slices with chloroquine, a compound that increases the pH of acidic subcellular organelles, for 10 h reduced the activity of cathepsin L by 83 +/- 0.87% (mean +/- s.e.m.) while only marginally suppressing cathepsin B. This effect was followed within 3 h by an increase in the concentration of mature, single-chain cathepsin D (up 61 +/- 28%). Selective depression of cathepsin L with N-CBZ-L-phenylalanyl-L-phenylalanine-diazomethylketone also resulted in increases in enzymatically active cathepsin D and the delayed appearance of a 29 kDa fragment of the tau protein. These findings demonstrate that the pattern of cathepsin L, B, and D changes found in the aged brain can be reproduced by reducing the acidity of the lysosomal milieu. They also indicate that such pH shifts initiate a sequence of linked disturbances (inactivation of cathepsin L > induction of cathepsin D > aberrant tau proteolysis) likely to play an important role in brain ageing.
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PMID:Selective suppression of cathepsin L results from elevations in lysosomal pH and is followed by proteolysis of tau protein. 967 99

Previous studies have shown that lysine residues on the surface of cathepsins and other lysosomal proteins are a shared component of the recognition structure involved in mannose phosphorylation. In this study, the involvement of specific lysine residues in mannose phosphorylation of cathepsin D was explored by site-directed mutagenesis. Mutation of two lysine residues in the mature portion of the protein, Lys-203 and Lys-293, cooperated to inhibit mannose phosphorylation by 70%. Other positively charged residues could not substitute for lysine at these positions, and comparison of thermal denaturation curves for the wild type and mutant proteins indicated that the inhibition could not be explained by alterations in protein folding. Structural comparisons of the two lysine residues with those required for phosphorylation of cathepsin L, using models generated from recently acquired crystal structures, revealed several relevant similarities. On both molecules, the lysine residues were positioned approximately 34 A apart (34.06 A for cathepsin D and 33.80 A for cathepsin L). When the lysine pairs were superimposed, N-linked glycosylation sites on the two proteins were found to be oriented so that oligosaccharides extending out from the sites could share a common region of space. Further similarities in the local environments of the critical lysines were also observed. These results provide details for a common lysosomal targeting structure based on a specific arrangement of lysine residues with respect to each other and to glycosylation sites on the surface of lysosomal proteins.
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PMID:Lysine-based structure responsible for selective mannose phosphorylation of cathepsin D and cathepsin L defines a common structural motif for lysosomal enzyme targeting. 969 59


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