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)

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

Salmonella typhimurium is an intracellular bacterial pathogen that remains enclosed in vacuoles (SCV) upon entry into the host cell. In this study we have examined the intracellular trafficking route of S. typhimurium within epithelial cells. Indirect immunofluorescence analysis showed that bacteria initiated fusion with lysosomal membrane glycoprotein (lgp)-containing compartments approximately 15 min after bacterial internalization. This process was completed approximately 75 min later and did not require microtubules. Cation-independent (CI)- or cation-dependent (CD)-mannose 6-phosphate receptors (M6PRs) were not observed at detectable levels in SCV. Lysosomal enzymes showed a different distribution in SCV: lysosomal-acid phosphatase (LAP) was incorporated into these vacuoles with the same kinetics as lgps, while cathepsin D was present in a low proportion (approximately 30%) of SCV. Uptake experiments with fluid endocytic tracers such as fluorescein-dextran sulphate (F-DX) or horseradish-peroxidase (HRP) showed that after 2 h of uptake, F-DX was present in approximately 75% of lgp-containing vesicles in uninfected cells, while only approximately 15% of SCV contained small amounts of the tracer during the same uptake period. SCV also showed only partial fusion with HRP-preloaded secondary lysosomes, with approximately 30% of SCV having detectable amounts of HRP at 6 h after infection. These results indicate that SCV show limited accessibility to fluid endocytic tracers and mature lysosomes, and are therefore functionally separated from the endocytic route. Moreover, the unusual intracellular trafficking route of S. typhimurium inside epithelial cells has allowed us to establish the existence of two different lgp-containing vesicles in Salmonella-infected cells: one population is separated from the endocytic route, fusogenic with incoming SCV and may arise from a secretory pathway, while the second involves the classical secondary or mature lysosomes.
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PMID:Targeting of Salmonella typhimurium to vesicles containing lysosomal membrane glycoproteins bypasses compartments with mannose 6-phosphate receptors. 769 96

Invasion of the cultured epithelial cell lines HeLa, Henle-407, and Caco-2 (polarized and nonpolarized) by Salmonella typhi and Salmonella typhimurium were compared using conventional gentamicin invasion assays. Additionally, the mechanisms of invasion and intracellular trafficking by S. typhi and S. typhimurium were compared in HeLa cells using indirect immunofluorescence microscopy. S. typhi strain Ty2 was invasive in all human cell lines tested, including apical uptake into polarized Caco-2 cell monolayers. This strain also replicated at levels similar to S. typhimurium strain SL1344 inside HeLa and Henle-407 cells. Indirect immunofluorescence microscopy confirmed that S. typhi, like S. typhimurium, induced membrane ruffles and cytoskeletal rearrangements upon contact with HeLa cell surfaces. Ruffling induced by S. typhi and S. typhimurium was accompanied by macropinocytosis of the fluid phase endocytic marker fluorescein-dextran-sulphate and by aggregation of cell surface class I MHC heavy chain. Intracellular lysosomal trafficking of S. typhi and S. typhimurium in HeLa cells was also studied. The lysosomal membrane glycoprotein marker h-lamp-2 colocalized with S. typhi-containing vacuoles, as previously shown for S. typhimurium. The soluble lysosomal enzyme marker cathepsin D also was found within S. typhi-containing vacuoles to the same extent as previously published for S. typhimurium. The results from this study suggest that S. typhi and S. typhimurium use similar mechanisms for invasion and intracellular trafficking in cultured human epithelial cells.
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PMID:Comparison of Salmonella typhi and Salmonella typhimurium invasion, intracellular growth and localization in cultured human epithelial cells. 775 82

We studied the biogenesis of the acrosome in sperm cells in immunogold-labeled ultrathin cryosections of rat testis, using a variety of antibodies against endosomal/lysosomal marker protein and acrosin, the major secretory protein of sperm cells. As expected, acrosomes and proacrosomal vesicles in the trans-Golgi region contained abundant acrosin. Rat lysosomal membrane glycoprotein (lgp) 120 and mouse lysosome-associated membrane protein-1 were not detectable in the acrosomal membrane. Similarly, the late endosomal markers cation-dependent and -independent mannose 6-phosphate receptors were absent from the acrosome and proacrosomal vesicles. Therefore, acrosomes do not exhibit these endosomal/lysosomal features. Apart from (pro) acrosomal vesicles, both spermatocytes and spermatids contained classical lysosomes (positive for rat lgp 120, mouse lysosome-associated membrane protein-1, and cathepsin D) that were negative for acrosin. Quantitative analysis of the immunogold labeling showed that spermatocytes express more mannose 6-phosphate receptors and lgp 120 than spermatids, whereas the opposite situation existed for acrosin. These data indicate differential synthetic activity of lysosomal and acrosomal constituents in different states of sperm differentiation. Together, our observations argue against a lysosomal /endosomal origin of the acrosome.
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PMID:Evidence for a nonlysosomal origin of the acrosome. 860 90

Stable BHK cell lines inducibly expressing wild-type or dominant negative mutant forms of the rab7 GTPase were isolated and used to analyze the role of a rab7-regulated pathway in lysosome biogenesis. Expression of mutant rab7N125I protein induced a dramatic redistribution of cation-independent mannose 6-phosphate receptor (CI-MPR) from its normal perinuclear localization to large peripheral endosomes. Under these circumstances approximately 50% of the total receptor and several lysosomal hydrolases cofractionated with light membranes containing early endosome and Golgi markers. Late endosomes and lysosomes were contained exclusively in well-separated, denser gradient fractions. Newly synthesized CI-MPR and cathepsin D were shown to traverse through an early endocytic compartment, and functional rab7 was crucial for delivery to later compartments. This observation was evidenced by the fact that 2 h after synthesis, both markers were more prevalent in fractions containing light membranes. In addition, both were sensitive to HRP-DAB- mediated cross-linking of early endosomal proteins, and the late endosomal processing of cathepsin D was impaired. Using similar criteria, the lysosomal membrane glycoprotein 120 was not found accumulated in an early endocytic compartment. The data are indicative of a post-Golgi divergence in the routes followed by different lysosome-directed molecules.
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PMID:Mutant Rab7 causes the accumulation of cathepsin D and cation-independent mannose 6-phosphate receptor in an early endocytic compartment. 949 Jul 21

A fluorescent antigen, FITC10BSA, that is sensitive to several of the biochemical processes involved in antigen processing was constructed. In combination with both flow cytometry and subcellular fractionation, the unique probe provided new details regarding the kinetics and intracellular pathways involved in antigen processing in murine macrophage. These studies suggested that macrophage utilized multiple vesicles as opposed to a few specific organelles for major histocompatibility complex (MHC) type II-peptide loading and transport. Although newly formed MHC II-peptide complexes were detected in cathepsin D-positive, lysosomal associated membrane glycoprotein (LAMP-1)-positive lysosomes, MHC II-peptide loading also occurred in transferrin receptor-positive endosomes. Interestingly, MHC II-fluoresceinated complexes were only observed in transferrin receptor-positive organelles as opposed to MHC II-unlabelled peptide complexes which were detected in traditional early lysosomal compartments. More importantly, MHC II-peptide complexes were monitored in light transferrin receptor-positive fractions following their initial appearance in dense endosomal/lysosomal fractions. Control experiments suggested that these complexes represented intermediates in the process of migrating to the cell surface through a retrograde pathway within the macrophage.
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PMID:Kinetics and intracellular pathways required for major histocompatibility complex II-peptide loading and surface expression of a fluorescent hapten-protein conjugate in murine macrophage. 1023 42