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:1.11.1.7 (peroxidase)
65,474 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The endocytic pathway of horseradish peroxidase (HRP) was investigated in the perikarya of cultured neurons by electron microscopy and enzyme cytochemistry. The tracer was observed in endocytic pits and vesicles, endosomes, multivesicular bodies, and lysosomes. It took approximate 15 min for the transfer of HRP from the exterior of the cell to the lysosomes. 2. Monensin induced distension of the Golgi apparatus and formation of intracellular vacuoles. When neurons were incubated with both monensin and HRP for 30 to 120 min, the number of HRP-labeled endosomes was greater than that in the monensin-free group, whereas the reverse was seen for HRP-positive lysosomes. The formation of HRP-positive lysosomes in monensin-treated cells was blocked by 47 to 79%. 3. These results indicate that the intracellular transport of the endocytosed macromolecule is pH dependent. It is also possible that the export of lysosomal enzymes is inhibited by monensin, resulting in an accumulation of the endosomes and a reduction of the lysosomes.
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PMID:Effect of monensin on the neuronal ultrastructure and endocytic pathway of macromolecules in cultured brain neurons. 139 68

A conjugate of horseradish peroxidase (HRP) to poly(L-lysine) (PLL) was used to characterize a non-lysosomal proteolytic compartment in the MDCK Strain I epithelial cell line. This compartment is expressed in a polar fashion, and is capable of degradation of the PLL moiety in the conjugate followed by release of HRP via a basal-to-apical, but not apical-to-basal, transcytotic pathway. This uptake, cleavage, and transport process appears to require approximately 2 hr, as there is a 2 hr lag-time between conjugate administration to the basal surface and HRP release to the apical medium. Monensin (10 microM) failed to inhibit this process, indicating that participation of the trans-Golgi network (TGN) in the trafficking of internalized conjugate is not the rate-determining step. Inhibition of HRP transport was found to be elicited by 50 micrograms/ml leupeptin, but only when applied to the basal surface. Brief trypsinization of either the basal or apical surfaces of cells preloaded with HRP conjugate showed no appreciable inhibitory effect on the apical release of HRP, indicating that an intracellular compartment rather than surface-bound enzymes is responsible for the degradation of the PLL moiety in the conjugate. Our results demonstrate the presence of an intracellular proteolytic compartment which is accessible in the basal-to-apical, but not apical-to-basal, transport pathway; and this compartment can be exploited for the transcytosis of membrane-bound molecules.
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PMID:Polarity in the transcytotic processing of apical and basal membrane-bound peroxidase-polylysine conjugates in MDCK cells. 173 33

The effect of monensin on endocytosis, transcytosis, recycling and transport to the Golgi apparatus in filter-grown Madin-Darby canine kidney (MDCK) cells was investigated using 125I-labeled ricin as a marker for membrane transport, and horseradish peroxidase (HRP) as a marker for fluid phase transport. Monensin (10 microM) stimulated transcytosis of both markers about 3-fold in the basolateral to apical direction. Transcytosis of HRP in the opposite direction, apical to basolateral, was reduced to approximately 50% of the control by monensin, whereas that of ricin was slightly increased. Recycling of markers endocytosed from the apical surface was reduced in the presence of monensin and there was an increased accumulation of both ricin and HRP in the cells. Transport of ricin to the Golgi apparatus increased to the same extent as the increase in intracellular accumulation. No change in recycling or accumulation was observed with monensin when the markers were added basolaterally, but transport of ricin to the Golgi apparatus increased almost 3-fold. Our results indicate that basolateral to apical transcytosis is increased in the absence of low endosomal pH, and they suggest that apical to basolateral transcytosis of a membrane-bound marker (ricin) is affected by monensin differently from that of a fluid phase marker (HRP).
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PMID:Effect of monensin on ricin and fluid phase transport in polarized MDCK cells. 179 Nov 87

A kinetic study of horseradish peroxidase transport was carried out on short-term cultured adult rat hepatocytes. After 4 hr of plating, cells were preincubated with monensin (1 microM) for 1 hr before their incubation with horseradish peroxidase for different times. Monensin treatment resulted in the dilatation of the Golgi apparatus and caused the appearance of numerous intracellular lumina lined with microvilli in associated as well as isolated hepatocytes but did not modify newly formed bile canaliculi. The frequency of their appearance increased to 28% in cells pretreated with monensin compared to 2% in controls. Intracellular lumina membranes had the morphological features of apical membranes and were stained with horseradish peroxidase more frequently than those of newly formed bile canaliculi. This work therefore provides a model for studying bile secretion in cultured hepatocytes. Our results also suggest that the biliary transport of horseradish peroxidase does not involve the Golgi apparatus, since horseradish peroxidase was never observed in the Golgi stacks even after monensin treatment.
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PMID:The effects of monensin on the transport of horseradish peroxidase into intracellular lumina in cultured rat hepatocytes. 273 4

The biosynthesis of myeloperoxidase in human promyelocytic leukemia HL-60 cells was studied by pulse-chase and immunoprecipitation methods and separation of subcellular organelles using Percoll density gradient fractionation. These studies revealed that in control and monensin (1 microM) treated cells, more than 85% of the total immunoprecipitable radiolabeled myeloperoxidase was present predominantly in precursor form (Mr 91,000) and resided in lower density compartments after an initial 3-h labeling period. Using biochemical and ultrastructural techniques, the lower density regions of the gradient were found to contain elements of the endoplasmic reticulum and the Golgi complex. Following a 16-h chase period, about 70% of the radiolabeled myeloperoxidase in untreated cells was found predominantly in denser regions of the gradient and was present mainly in the form of the mature large subunit (Mr 63,000). These dense regions were shown to contain azurophilic granules by means of the distribution of beta-glucuronidase and myeloperoxidase activities and by electron microscopy. Processing of myeloperoxidase and its deposition into dense granules were blocked by monensin treatment. Following a 16-h chase period in the presence of monensin, approximately 80% of the radiolabeled myeloperoxidase continued to reside in lower density compartments and was predominantly in precursor (Mr 91,000) and intermediate (Mr 81,000 and 74,000) forms. Only about 10% of the radiolabeled myeloperoxidase was associated with dense azurophilic granules. Monensin treatment produced large, Golgi-derived vacuoles which were isolated using Percoll density centrifugation and identified by electron microscopy. These vacuoles were found to be essentially devoid of peroxidase activity and pulse-labeled, newly synthesized radiolabeled myeloperoxidase species. The effects of monensin on transport and processing were reversible after a 3-h exposure and 16-h chase period in the absence of monensin. Taken together, these data indicate that maturation of myeloperoxidase is closely linked to its deposition into dense azurophilic granules via a monensin-sensitive process(es). The lower density compartments within which immature myeloperoxidase species accumulate in the presence of monensin appear to be functionally related to or associated with Golgi or endoplasmic reticulum structures distinct from the large monensin-induced vacuoles.
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PMID:Biochemical and ultrastructural effects of monensin on the processing, intracellular transport, and packaging of myeloperoxidase into low and high density compartments of human leukemia (HL-60) cells. 282 13

The processing and intracellular transport of lactoferrin of the neutrophil specific granules was investigated by biosynthetic labeling with (14C)leucine of bone marrow cells from healthy individuals and patients with chronic myeloid leukemia. Lactoferrin was precipitated with antilactoferrin serum and the immunoprecipitates were analyzed by sodium dodecyl sulfate (SDS), polyacrylamide gel electrophoresis (PAGE) followed by fluorography. In contrast to myeloperoxidase of azurophil granules, lactoferrin was not synthesized as a larger precursor, and it was not found to be phosphorylated. The transfer to granules of newly synthesized lactoferrin was demonstrated in pulse-chase labeling experiments followed by centrifugation of cell homogenate in a Percoll gradient. Monensin, which exchanges protons for Na+ and NH4+ cation, blocked the transfer completely, indicating a need for acidification mechanisms. Unlike myeloperoxidase, newly synthesized lactoferrin rapidly became resistant to endoglycosidase H, indicating a transport through the medial and transcisternae of the Golgi apparatus with conversion of "high mannose" to "complex" oligosaccharide side chains. Intracellular transfer of some major neutrophil azurophil and specific granule constituents is obviously regulated differently. Lactoferrin seems to be processed like proteins destined for secretion, while myeloperoxidase is processed more or less like lysosomal enzymes.
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PMID:Biosynthesis and processing of lactoferrin in bone marrow cells, a comparison with processing of myeloperoxidase. 282 14

Human eosinophil peroxidase (EPO) was purified from leukocytes obtained from a patient with hypereosinophilia. EPO was extracted from the granule fraction using 0.2 mol/L sodium acetate pH 4.0, and the extract was subjected to gel chromatography on Sephadex G-75 and ion exchange chromatography on Biorex 70. The mol wt calculated from gel chromatography was approximately 50,000. However, under reducing and denaturing conditions, polyacrylamide gel electrophoresis revealed two subunits with mol wt of 50,000 and 15,000. The biosynthesis of EPO was studied in marrow cells from patients with eosinophilia using labeling with (14C)-leucine, followed by immunoprecipitation with anti-EPO, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and fluorography for visualization of labeled EPO. Biosynthesis of an Mr 53,000 subunit was demonstrated. Biosynthetic labeling of the Mr 15,000 subunit was not demonstrated. A labeled Mr 25,000 chain was detected and may represent a degradation product or a chain that, after further modification, produces the Mr 15,000 subunit. Labeling was also detected in two polypeptides with mol wt of 78,000 and 72,000. These forms of EPO seem to represent precursor polypeptides subjected to proteolytic processing in a similar manner as has been reported for myeloperoxidase (MPO). However, Monensin, a proton ionophore, which blocks the processing of MPO, did not inhibit processing of EPO, indicating separate mechanisms by which MPO and EPO are directed to granules.
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PMID:Purification of eosinophil peroxidase and studies of biosynthesis and processing in human marrow cells. 299 61

Myeloperoxidase, stored in azurophil granules of neutrophils, is synthesized in promyelocytes as a larger molecular weight precursor, which is processed to yield a transient Mr 82 000 intermediate and mature polypeptides with molecular weights of 62 000 and 12 000. We have tried to define subcellular sites for processing using metabolic labelling of the promyelocytic leukemia cell line HL-60 in combination with subcellular fractionation on a Percoll gradient. A reasonable separation was achieved between azurophil granules, Golgi elements and endoplasmic reticulum. The finding of almost exclusively fully processed myeloperoxidase in granules and a mixture of unprocessed and processed polypeptide in fractions enriched in Golgi elements suggests that processing occurred mainly in pregranular structures. Monensin, which exchanges protons for Na+, and the base chloroquine blocked processing probably by inhibition of transport through the Golgi apparatus. However, the lysosomotropic NH4+ cation did not inhibit processing or transport indicating that processing is not necessarily influenced by pH-dependent mechanisms. Results from digestion with endoglycosidase H, incubation with tunicamycin and metabolic labelling with [3H]mannose indicated that myeloperoxidase contained high mannose oligosaccharide side chains. Also [32P]phosphate incorporated into Mr 90 000 and Mr 62 000 myeloperoxidase was susceptible to endoglycosidase H indicating that oligosaccharide side chains are modified by phosphorylation as in lysosomal enzymes. Thus, even if myeloperoxidase contained mannose 6-phosphate residues, these may not necessarily be involved in directing transport to the azurophil granules.
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PMID:The processing and intracellular transport of myeloperoxidase. Modulation by lysosomotropic agents and monensin. 300 51

Single cell suspensions from 15-day embryonic rat hindbrain plated on collagen formed large clumps by day 1 in culture. Neurite outgrowth was visible within 2 days. By day 14, morphological synapses were observed in nearly all instances of contact of a neurite ending with another cell. At day 3 in culture, the Golgi apparatus consisted of relatively few, broad lamellae. By contrast, at day 7 in culture this organelle consisted of tightly packed lamellar stacks with a considerable increase in vesicles budding from lamellae. Electron-lucent vesicles, ranging in size from 60 to 180 nm, similar to those generated by the Golgi apparatus were noted in neurite shafts and growth cones, with fusion of these vesicles virtually exclusively at the growth cone leading edge. Monensin resulted in the loss of these vesicles in cell somata and neuritic profiles. The electron-dense marker horseradish peroxidase was not incorporated into these vesicles following its addition to the culture medium, indicating that the vesicles were exocytotic. The number of total vesicles increased during the first 7 days of neurite outgrowth with no further increase up to day 14. This increase was due entirely to vesicles not labeled with the impermeable electron-dense stain ruthenium red, indicating that this increase represents actual vesicular elements and not increased surface convolutions. These data suggest that the 60- to 180-nm electron lucent vesicles are derived from the Golgi apparatus and, by fusion with the growth cone plasmalemma, provide new membrane required for neuritic outgrowth and maintenance.
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PMID:Vesicle-mediated delivery of membrane to growth cones during neuritogenesis in embryonic rat primary neuronal cultures. 318 97

The biosynthesis and carbohydrate processing of the insulin receptor were studied in cultured human lymphocytes by means of metabolic and cell surface labeling, immunoprecipitation with anti-receptor autoantibodies, and analysis on sodium dodecyl sulfate-polyacrylamide gels under reducing conditions. In addition to the two major subunits of Mr = 135,000 and Mr = 95,000, two higher molecular weight bands were detected of Mr = 210,000 and Mr = 190,000. The Mr = 210,000 band and the two major subunits were labeled by [3H]mannose, [3H]glucosamine, [3H]galactose, and [3H]fucose, and were bound by immobilized lentil, wheat germ, and ricin I lectins. On the other hand, the Mr = 190,000 band was labeled only by [3H]mannose and [3H]glucosamine and was bound only by lentil lectin. All four components could be labeled with [35S] methionine; however, in contrast with the other three polypeptides, the Mr = 190,000 band was not labeled by cell surface iodination with lactoperoxidase, suggesting that it is not exposed at the outer surface of the plasma membrane. Pulse-chase studies with [3H]mannose showed that the Mr = 190,000 was the earliest labeled component of the receptor; radioactivity in this band reached a maximum 1 h after the pulse, clearly preceded the appearance of the other components, and had a very brief half-life (t1/2 = 2.5 h). The Mr = 210,000, Mr = 135,000, and Mr = 95,000 bands were next in appearance and reached a maximum 6 h in the chase period. Monensin, an ionophore which interferes with maturation of some proteins, blocked both the disappearance of the Mr = 190,000 protein and the appearance of the Mr = 135,000 and Mr = 95,000 subunits. The mannose incorporated in the Mr = 190,000 component was fully sensitive to treatment with endoglycosidase H while that in the Mr = 210,000 band and the two major subunits was only partially sensitive. Tryptic fingerprints of the 125I-labeled Mr = 210,000 band suggested that this component contains peptides of both the Mr = 135,000 and Mr = 95,000 subunits. In conclusion, the Mr = 190,000 component appears to represent the high mannose precursor form of the insulin receptor that undergoes carbohydrate processing and proteolytic cleavage to generate the two major subunits. In addition, the Mr = 210,000 band is probably the fully glycosylated form of the precursor that escapes cleavage and is expressed in the plasma membrane.
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PMID:Biosynthesis and glycosylation of the insulin receptor. Evidence for a single polypeptide precursor of the two major subunits. 641


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