Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.9.3.1 (cytochrome oxidase)
 8,822 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

As a first step in studies of the molecular mechanism(s) underlying gentamicin toxicity, rat kidney cortex has been subfractionated using differential centrifugation. An analytical, rather than preparative approach was used. DNA was used as a marker for the nuclei, cytochrome oxidase for mitochondria, acid phosphatase for lysosomes, catalase for peroxisomes, NADPH-cytochrome c reductase for the endoplasmic reticulum, p-nitrophenyl-alpha-mannosidase (at pH 5.5) for the Golgi apparatus, AMPase for the plasma membrane in general, and alkaline phosphatase for the brush border, and lactate dehydrogenase for the cytosol. In addition, electron microscopy was performed on the subfractions obtained. The distributions of subcellular markers obtained here for the rat kidney cortex closely resemble the corresponding distributions reported for rat liver. This procedure can now be used to look for biochemical and/or toxic changes which might be reflected in an altered distribution pattern for marker enzymes.
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PMID:Biochemical effects of gentamicin on rat kidney cortex. I. Analytical subfractionation of control tissue. 303 Jul 99

As a first step in studies on the molecular mechanism(s) underlying gentamicin toxicity, the effect of treating rats with this aminoglycoside antibiotic (100 mg/kg once or twice daily for 3 days) on the analytical subfractionation of the kidney cortex has been examined. DNA was used as a marker for the nuclei, cytochrome oxidase for mitochondria, acid phosphatase for lysosomes, catalase for peroxisomes (with reservations; see the companion paper), NADPH-cytochrome c reductase for the endoplasmic reticulum, p-nitrophenyl-alpha-mannosidase (at pH 5.5) for the Golgi apparatus, AMPase for the plasma membrane in general and alkaline phosphatase for the brush border, and lactate dehydrogenase for the cytosol. In addition, the presumptive lysosomal hydrolases N-acetyl-beta-D-glucosaminidase, p-nitrophenyl-alpha-mannosidase (at pH 4.5), cathepsin D, and DNase II were monitored. Electron microscopy was also performed on the subfractions obtained. The only significant biochemical changes brought about by gentamicin treatment were that N-acetyl-beta-D-glucosaminidase demonstrated both a greater total activity and a larger enrichment in the 104,000gav pellet, while p-nitrophenyl-alpha-mannosidase at pH 4.5 demonstrated the same total activity and a greater enrichment in the 104,000gav pellet. Since myeloid bodies were shown by electron microscopy to sediment primarily with the 500gav and 10,000gav pellets, the biochemical changes seen cannot be associated with these morphological structures. These findings suggest that selective changes in a certain subpopulation(s) of lysosomes or in certain lysosomal enzymes may be involved in the early stages of gentamicin toxicity. On the other hand, no lysosomal membrane damage was observed here, since both the latency of acid phosphatase and the recovery of this activity in the soluble cytosol were unchanged. The present investigation may also have relevance for the dosage and duration of gentamicin treatment chosen in clinical situations.
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PMID:Biochemical effects of gentamicin on rat kidney cortex. II. Analytical subfractionation after short-term, high-dose treatment. 303 Aug

Homogenates of control and diet-induced atherosclerotic aortas of rabbit were prepared and the levels of DNA, protein, free and esterified cholesterol, and six enzymes known to be associated with various subcellular organelles [N-acetyl-beta-glucosaminidase, beta-galactosidase (lysosomes); cytochrome oxidase (mitochondria); neutral alpha-glucosidase (endoplasmic reticulum); 5'-nucleotidase (plasma membrane); catalase (peroxisomes)] were compared between control and atherosclerotic preparations. The levels of prostaglandins I2, E2, and F2 alpha, based on DNA, also were measured by radioimmunoassay. Atherosclerotic aortas were significantly enriched in catalase activity (440%) and in each of the acid hydrolases (395 and 630%), based on DNA, as well as in free (630%) and esterified cholesterol (930%), based on tissue wet weight, compared to control aortas. The control level of prostaglandin I2 was 10-fold higher than that of prostaglandin E2, which was 3-fold higher than that of prostaglandin F 2 alpha. Prostaglandin I2 doubled in amount with advanced atherosclerosis, while prostaglandin E2 increased over 10-fold, resulting in twice the amount of prostaglandin I2 than E2 in advanced atherosclerosis; the level of prostaglandin F2 alpha did not appear to change significantly with atherosclerosis. Increased levels of prostaglandins I2 and E2 were correlated significantly with increased aortic total cholesterol content (based on DNA) but not increased serum cholesterol levels. N-Acetyl-beta-glucosaminidase activity also was correlated significantly to aortic total cholesterol content and beta-galactosidase activity, as well as to the level of prostaglandin I2; in contrast, N-acetyl-beta-glucosaminidase was not significantly correlated to prostaglandin E2. The association of prostaglandins I2 and E2 with aortic total cholesterol suggests the participation of prostaglandins in the response of arterial cells to lipid accumulation in atherosclerosis. The specific association of aortic prostaglandin I2 level and N-acetyl-beta-glucosaminidase activity further suggests a possible role for this prostaglandin during arterial intralysosomal cholesterol accumulation.
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PMID:Arterial prostaglandins and lysosomal function during atherogenesis. I. Homogenates of diet-induced atherosclerotic aortas of rabbit. 389 3

A new method for the subcellular and cytochemical demonstration of cytochrome oxidase has been developed with the introduction of N-benzyl-p-phenylenediamine (BPDA) and the discovery that indoanilines are osmiophilic. These indoanilines produced upon oxidation of BPDA in the presence of naphthols are highly colored compounds that yield electron-opaque coordination polymers of osmium (osmium black) that are amorphous, insoluble in water, and in organic solvents. The best methods for preparing rat tissue were in decreasing order: fixation in formaldehyde solution, fresh tissue slices, and frozen sections of fresh or fixed tissue. Ultrathin sections were counterstained by bridging with the thiocarbohydrazide-osmium tetroxide (T-O) procedure for enhancing underlying membranous structures. Cytochrome oxidase activity was noted primarily in mitochondria and occasionally in sarcotubules of heart, in mitochondria and occasionally in infoldings of the plasma membrane of renal tubular cells, and in mitochondria and, to a great extent, in endoplasmic reticulum of hepatic cells. Cytochrome oxidase activity produced deposits in droplet form, whereas dehydrogenase activity resulted in uniform staining of mitochondrial cristae, as recently demonstrated with an osmiophilic tetrazolium salt. Even more recently we have succeeded in demonstrating cytochrome oxidase activity in nondroplet staining on mitochondrial cristae with an osmiophilic benzidine-type reagent that apparently polymerizes upon oxidation (to be published later).
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PMID:Ultrastructural demonstration of cytochrome oxidase activity by the Nadi reaction with osmiophilic reagents. 429 7

Improved, largely automated methods are described for the purification and analysis o peroxisomes, lysosomes, and mitochondria from the livers of rats injected with Triton WR-1339. With these new methods, it has become possible to obtain, in less than 6 hr and with reliable reproducibility, mitochondria practically free of contaminants, as well as the rarer cytoplasmic particles in amounts (about 100 mg of protein) and in a state of purity (95%) that make them suitable for detailed biochemical studies. The results obtained so far on these preparations have made more conclusive and precise previous estimates of the biochemical and morphological properties of the three groups of cytoplasmic particles. In addition, peroxisomes were found to contain essentially all the L-alpha-hydroxy acid oxidase of the liver, as well as a small, but significant fraction of its NADP-linked isocitrate dehydrogenase activity. Another small fraction of the latter enzyme is present in the mitochondria, the remainder being associated with the cell sap. The mitochondrial localization of the metabolically active cytoplasmic DNA could be verified. The relative content of the fractions in mitochondria, whole peroxisomes, peroxisome cores, lysosomes, and endoplasmic reticulum was estimated independently by direct measurements on electron micrographs, and by linear programming (based on the assumption that the particles are biochemically homogeneous) of the results of enzyme assays. The two types of estimates agreed very well, except for one fraction in which low cytochrome oxidase activity was associated with mitochondrial damage.
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PMID:The large-scale separation of peroxisomes, mitochondria, and lysosomes from the livers of rats injected with triton WR-1339. Improved isolation procedures, automated analysis, biochemical and morphological properties of fractions. 429 86

Enzymatic activities associated with Golgi apparatus-, endoplasmic reticulum-, plasma membrane-, mitochondria-, and microbody-rich cell fractions isolated from rat liver were determined and used as a basis for estimating fraction purity. Succinic dehydrogenase and cytochrome oxidase (mitochondria) activities were low in the Golgi apparatus-rich fraction. On the basis of glucose-6-phosphatase (endoplasmic reticulum) and 5'-nucleotidase (plasma membrane) activities, the Golgi apparatus-rich fraction obtained directly from sucrose gradients was estimated to contain no more than 10% endoplasmic reticulum- and 11% plasma membrane-derived material. Total protein contribution of endoplasmic reticulum, mitochondria, plasma membrane, microbodies (uric acid oxidase), and lysosomes (acid phosphatase) to the Golgi apparatus-rich fraction was estimated to be no more than 20-30% and decreased to less than 10% with further washing. The results show that purified Golgi apparatus fractions isolated routinely may exceed 80% Golgi apparatus-derived material. Nucleoside di- and triphosphatase activities were enriched 2-3-fold in the Golgi apparatus fraction relative to the total homogenate, and of a total of more than 25 enzyme-substrate combinations reported, only thiamine pyrophosphatase showed a significantly greater enrichment.
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PMID:Isolation of a Golgi apparatus-rich fraction from rat liver. II. Enzymatic characterization and comparison with other cell fractions. 431 70

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

1. A lag period of about 4 days preceded the onset of metamorphosis precociously induced by tri-iodothyronine in tadpoles of the giant American bullfrog (Rana catesbeiana). It was established by the accelerated synthesis or induction of carbamoyl phosphate synthetase and cytochrome oxidase in the liver, serum albumin and adult haemoglobin in the blood, acid phosphatase in the tail, and the increase in the hindleg/tail length ratio. 2. A 4- to 6-fold stimulation, 2 days after the induction of metamorphosis, of the rate of synthesis of rapidly labelled nuclear RNA in liver cells was followed by an increasing amount of RNA appearing in the cytoplasm. Most of the newly formed RNA on induction of metamorphosis was of the ribosomal type. An accelerated turnover at early stages of development preceded a net accumulation of RNA in the cytoplasm, with no change in the amount of DNA per liver. 3. Most hepatic ribosomes of the pre-metamorphic tadpoles were present as 78s monomers and 100s dimers; metamorphosis caused a shift towards larger polysomal aggregates with newly formed ribosomes that were relatively more tightly bound to membranes of the endoplasmic reticulum. 4. The appearance of new polyribosomes in the cytoplasm on induction of metamorphosis was co-ordinated in time with a stimulation of synthesis of phospholipids of the smooth and rough endoplasmic reticulum, followed by a gradual shift in preponderance from the smooth to the rough type of microsomal membranes. 5. Electron- and optical-microscopic examination of intact hepatocytes revealed a striking change in the distribution and nature of ribosomes and microsomal membranes during metamorphosis. 6. Ribosomes prepared from non-metamorphosing and metamorphosing animals were identical in their sedimentation coefficients and in the structural ribosomal proteins. The base composition and sedimentation coefficients of ribosomal RNA were also identical. Induction of metamorphosis also did not alter the incorporation of (32)P into the different phospholipid constituents of microsomal membranes. 7. Nascent (14)C-labelled protein with the highest specific activity was recovered in the ;heavy' rough membrane fraction of microsomes, whereas little (14)C was associated with ;free' polysomes. Protein synthesis in vivo was most markedly stimulated during metamorphosis in the tightly membrane-bound ribosomal fraction after the appearance of new ribosomes. 8. The rate of synthesis of macromolecules in vivo could not be followed beyond 7-8 days after induction because of variable shifts in precursor pools due to regression of larval tissues. 9. The stimulation of RNA and ribosome formation was specifically associated with the process of metamorphosis since no similar response to thyroid hormones occurred in those species (Axolotl and Necturus) in which the hormones failed to induce metamorphosis.
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PMID:The formation, distribution and function of ribosomes and microsomal membranes during induced amphibian metamorphosis. 558 18

The localization of the anion-sensitive ATPase (EC 3.6.1.3) of bovine corneal endothelium has been investigated. Homogenates were fractionated by differential and density gradient centrifugation, into fractions enriched in plasma membranes and mitochondria. (Na+ + K+)-ATPase (EC 3.6.1.3) and cytochrome oxidase (EC 1.9.3.1) were used as marker enzymes for these two cell components, and glucose-6-phosphatase (EC 3.1.3.5) was used to identify endoplasmic reticulum. 5'-Nucleotidase (EC 3.1.3.5) was also measured but was found not to be exclusively associated with any one cell component. The activity of the anion-sensitive ATPase (HCO3--ATPase) was measured in suspensions that were frozen and thawed before assay in order to expose latent enzyme activity. The fraction containing the greatest amount of (Na+ + K+)-ATPase (35%) contained only 6% of the cytochrome oxidase and HCO3--ATPase. Conversely, the mitochondrial fraction, containing 40% of the cytochrome oxidase, contained about 40% of the HCO3--ATPase, but only 7% of the (Na+ + K+)-ATPase. The recoveries and relative degree of purification of the cytochrome oxidase and HCO3--ATPase were also nearly identical in the other fractions examined. It was concluded that the anion-sensitive ATPase activity of the corneal endothelium is located solely in the mitochondria and not in the plasma membrane. Consequently, any role that the enzymes may have in the transport of bicarbonate across this tissue, which had been suggested in earlier studies, must be an indirect one.
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PMID:The localization of the anion-sensitive ATPase activity in corneal endothelium. 611 46

In rat liver, three different enzymes with peroxidatic activity are demonstrated with modifications of the DAB-technique: peroxidase in the endoplasmic reticulum of Kupffer cells, catalase in peroxisomes and cytochrome oxidase in mitochondria. The major problem of the DAB-methods is their limited specificity so that often in tissues incubated for one enzyme the other two proteins are also stained simultaneously. We have studied the conditions for selective staining of each of these three enzymes in rat liver fixed either by perfusion with glutaraldehyde or by immersion in a modified Karnovsky's glutaraldehyde-formaldehyde fixative. The observations indicate that in perfusion fixed material selective staining can be obtained by reduction of the incubation time (5 min) and the use of optimal conditions for each enzyme. In livers fixed by immersion the distribution of the staining is patchy and irregular and usually longer incubation times (15-30 min) are required. Selective staining of peroxidase in Kupffer cells was obtained by brief incubation at room temperature in a medium containing 2.5 mM DAB in cacodylte buffer pH 6.5 and 0.02% H2O2. The exclusive staining for cytochrome oxidase in cristae of mitochondria was achieved after short incubation in 2.5 mM DAB in phosphate buffer pH 7.2 containing 0.05% cytochrome c. For selective demonstration of catalase in peroxisomes the tissue was incubated in 5 mM DAB in Teorell-Stenhagen (or glycine-NaOH) buffer at pH 10.5 and 0.15% H2O2. The prolongation of the incubation time in peroxidase medium caused marked staining of both mitochondria and peroxisomes. In the cytochrome oxidase medium longer incubations led to slight staining of peroxisomes. The catalase medium was quite selective for this enzyme so that even after incubation for 120 min only peroxisomes stained.
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PMID:Selective cytochemical localization of peroxidase, cytochrome oxidase and catalase in rat liver with 3,3'-diaminobenzidine. 626 82


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