Acid Deoxyribonuclease Activity Research Articles

Lysosomal deoxyribonuclease II from the mussel Mytilus galloprovincialis: Characterization and seasonal activity

In view of the importance of Mytilus species in marine ecology and environmental quality assessment, all aspects of mussel enzymes that are related to environmental factors are of great relevance. A lysosomal enzyme, deoxyribonuclease II, a member of the acid deoxyribonucleases, has a major role in cellular DNA degradation. The presence and seasonal variations of the acid deoxyribonuclease (DNase) activity in the digestive gland and gills of the mussel Mytilus galloprovincialis were investigated. The enzyme was localized in the lysosomal vacuolar system of both gill and digestive gland tissues. Two active acid DNase isoforms were detected in the digestive gland (48 and 37 kDa) and one in the gills (48 kDa). All active enzyme isoforms coincided with protein bands immunoreactive to antibodies raised against human DNase II. Post-translational lysosomal processing of acid DNase in the digestive gland but not in the gills indicated tissue-specific enzyme maturation. The activity of acid DNase determined across the annual cycle showed a tissue-specific pattern that could be related to the seasonal activity of mussel organs and their biological function. In the digestive gland, high DNase activity was observed in relation to food availability and spawning. Higher enzyme activity in the gills observed in the autumn/winter period could be associated with feeding and cellular adaptation to low seawater temperatures.

Picrotoxin-induced convulsions and lysosomal function in the rat brain

The effect of picrotoxin-induced convulsions on lysosomal function in rat brain were evaluated by measuring the free as well as total acid phosphatase, cathepsin D, acid ribonuclease (RNAse II) and acid deoxyribonuclease (DNAse II) activities. Following picrotoxin treatment the free RNAse II activity increased whereas the total activities of practically all the other enzymes decreased. Paradoxically, the cathepsin D activity, free as well the total was completely abolished. In case of all the enzymes the ratio of Total activity/Free activity decreased indicating increased lysosomal membrane fragility which could lead to process of neurodegeneration in the epileptic animals.

Activity of acid deoxyribonuclease towards diastereoisomers of thymidyl 3'-(4-nitrophenyl phosphorothioate). Stereochemistry of transnucleotidylation reaction.

The (Rp)- and (Sp)-diastereoisomers of thymidyl 3'-(4-nitrophenyl phosphorothioate) (1) were found to act as unusual substrates for acid deoxyribonuclease (DNase II). Instead of the expected thymidine 3'-phosphorothioate, the product resulting from the reaction of (Rp)-1 catalyzed by DNase II was identified as (Sp, Rp)-thymidyl (3'-5')thymidyl phosphorothioate 3'-(4-nitrophenyl phosphorothioate), while that from (Sp)-1 has been recognized as a 10:1 mixture of (Sp, Rp)-thymidyl (3'-5')thymidyl phosphorothioate 5'-(4-nitrophenyl phosphorothioate) and (Rp, Sp)-thymidyl (3'-5')-thymidyl phosphorothioate 3'-(4-nitrophenyl phosphorothioate), respectively. Both types of transnucleotidylations were found to occur with retention of configuration at phosphorus. Stereochemical results may be interpreted in terms of two step mechanisms involving the formation of the intermediate, covalent substrate enzyme complexes.

慢性関節リウマチ関節液中の酸性ＤＮＡ分解酵素について （第一報）

An acid deoxyribonuclease activity was detected in synovial fluids of patients with rheumatoid arthritis (RA) and osteoarthritis (OA) by sensitive method using 32P-labeled Escherichia coli DNA as substrate for enzyme assay. Nineteen patients, 20 joints with RA and 7 patients, 10 joints with OA were investigated. The enzyme activity was dected in only 3 of 9 cases with RA classified in stage I and II of Steinbrcker's stage classification. The enzyme activity was detected in 4 of 5 cases with stage III and all of 6 cases with stage IV, most cases of which showed high enzyme activity. The higher enzyme activity in synovial fluids was observed in the cases with more advanced X-ray stage.

Electron cytochemical evidence for lysosomal-like equivalents in Histoplasma capsulatum.

Electron cytochemical localizations of acid phosphatase, aryl sulfatase, deoxyribonuclease, adenylate cyclase, and c-AMP phosphodiesterase activity sites in thin sections of cells of the two growth phases of the zoopathogenic Histoplasma capsulatum are described and illustrated by transmission electron micrographs. Various activity sites of these enzymes included the cytomembranes of the nucleus, mitochondria, and endoplasmic reticulum. At the same time, electron opaque reaction products were sequestered within membrane-bound, vacuolar regions of the cytosol. These vacuoles may be ontogenically related to membranous or vesicular inclusions commonly seen in thin sections of glutaraldehyde osmium tetroxide-fixed cells. These enzymatically-active vacuoles are believed consistent with previous descriptions of fungal lysosomal-like structures found in certain other fungi. Lysosomal-like vacuoles of H. capsulatum may provide a means of compartmentalization of various hydrolytic enzymes involved in catabolism and mobilization of storage reserves, and perhaps to function as well in other aspects of the life cycle of this important pathogenic dimorphic fungus.

Effect of antigenic stimulation on acid deoxyribonuclease activity in lymphocytes: I. Antibody-induced response

The effect of antigen-induced stimulation on acid deoxyribonuclease (DNase) activity in BALB/c mouse lymphoid cells was determined. Increase in acid DNase activity was found in intact spleen cell populations of mice from the second or fourth day after immunization with pneumococcal polysaccharide type III and from the fourth day after immunization with SRBC. DNase determinations performed with spleen cell fractions prepared from SRBC-immunized mice, showed that the rise in the enzyme activity was confined to the fraction containing the antibody-forming cells. The DNase activity was also increased in spleen cell cultures, stimulated with SRBC in vitro. Rise in the activity of this enzyme was also observed in peritoneal cell populations taken from SRBC-immunized mice. This change was maximal on the second day after immunization, when no appreciable increase in DNase activity of spleen cells was yet detected. The results obtained suggest, that acid DNase is an enzyme involved in the proliferative/maturation response to antigenic stimulation. It is a consequence of antigenic stimulation rather than being involved in the process of afferent stimulation.

Evaluation of the validity of the histochemical lead nitrate technique for alkaline and acid deoxyribonuclease.

The results of several tests and the characteristic morphological distribution of the enzymatic activity appeared to be in favor of the validity and specificity of the histochemical lead nitrate technique for alkaline and acid deoxyribonuclease (DNAse) detection. These tests included thermal inhibition, omission of substrate, use of different chemical inhibitors and the reproduction of histochemical staining on Coujard's slides. Most of these results were in conformity with the biochemical data gathered from literature. Topographically selective inhibition of alkaline or acid DNAse by different factors suggested that there might exist two kinds of alkaline or acid DNase--one cytoplasmic and the other one nuclear. The whole histochemical procedure produced relatively small loss of alkaline and acid DNAse activities as verified by biochemical methods.

Deoxyribonucleases in phytohemagglutinin-stimulated lymphocytes

The in situ assay of deoxyribonucleases in DNA-containing polyacrylamide gels following their separation by microdisc electrophoresis was used to determine the deoxyribonuclease pattern of human lymphocytes during stimulation with phytohemagglutinin (PHA). Two additional neutral deoxyribonuclease activities are detectable in stimulated cells, one only active with denatured DNA, the other active with native and denatured DNA as substrate, showing a maximum activity after 36 h and increasing in waves respectively. A group of acid deoxyribonuclease activities also shows a maximum after 36 h of stimulation. A neutral deoxyribonuclease active only with native DNA is missing in stimulated lymphocytes. It is suggested that the acid deoxyribonuclease activities and the neutral deoxyribonuclease active only with denatured DNA are involved in DNA synthesis, whereas the involvement of the neutral deoxyribonuclease active with native and denatured DNA in processing of DNA excreted in stimulated lymphocytes is discussed.

Effect of cholesterol on multiplication, lipid metabolism and lysosomal enzymes ofHartmanella culbertsoni

Addition of sonicated dispersions of cholesterol to peptone-salt-vitamin medium resulted in the metabolism of the sterol byHartmanella culbertsoni. Trophozoite multiplication was stimulated at 1–5 mg/litre, but retarded at 10–20 mg/litre. When cholesterol was added to the medium, incorporation of [1,2-14C] -acetate into neutral lipid, phospholipid, non-saponifiable and cholesterol fractions of the amoebae was significantly reduced. Cholesterol ester was detected in the medium but phospholipids were not released. Addition of cholesterol stimulated the activity of lysosomal acid phosphatase, acid deoxyribonuclease and cathepsin B but did not affect 5′-nucleotidase, adenosine triphosphatase, alkaline phosphatase, glucose-6-phosphatase, succinate dehydrogenase and cytochrome C oxidase.

Distribution of acid and alkaline deoxyribonucleases in white and grey matter regions of growing and aging chick brain

The activities of acid and alkaline deoxyribonucleases in the white and grey matter areas of growing and old chick cerebrum were measured. Two marker enzymes for glial cells, butyrylcholinesterase and carbonic anhydrase were also measured in these regions. Higher specific activities of both butyrylcholinesterase and carbonic anhydrase were found in the white matter region at all the stages studied. Acid and alkaline deoxyribonuclease activities were observed in both white and grey matter. The decrease in the specific activity of acid deoxyribonuclease with advancement of age was more pronounced as compared to the alkaline deoxyribonuclease Marked reduction in total acid deoxyribonuclease activity in white matter, beyond the age of 130 days, was observed. On the other hand, total alkaline deoxyribonuclease activity in both white and grey matter continued to increase with age Further, the activity per mg of DNA also increased in white matter of the old brain. These results indirectly suggest a continued role for alkaline deoxyribonuclease in glial cells formed at a later age.

Deoxyribonucleic acid polymerase and deoxyribonuclease activities in pea seedlings [proceedings

Deoxyribonucleic acid polymerase and deoxyribonuclease activities in pea seedlings [proceedings

Acid Hydrolase Activity During Growth and Encystment in Acanthamoeba castellanii*

The activity and sedimentation of acid phosphatase (APase), acid deoxyribonuclease (DNase), and acid ribonuclease (RNase) were investigated throughout growth and encystment in Acanthamoeba castellanii. The activities/mg protein of all 3 hydrolases are high in young cultures and decrease to constant levels in postlog cells. The RNase activity/ameba decreases 50% during growth, whereas the activity/cell of both APase and DNase remains constant. The percent sedimentation at 20,000 g of all 3 enzymes gradually increases from about 40% in midlog to a plateau of 80% in postlog cells. During encystment, the sedimentation behavior of RNase differs from that of APase and DNase. Encystment is characterized by a differential decrease in the activity/cell of the 3 hydrolases, with RNase decreasing most rapidly and APase least rapidly. APase is unique in that a transient increase of its specific activity is noted during encystment, even though its activity/cell is decreasing.

Notizen: Unterschiedliche Desoxyribonuclease-Aktivitäten im Seminalplasma von Bullen/Different Deoxyribonuclease-Activities in Bull Seminal Plasma

By means of the in situ assay of deoxyribonucleases in DNA-containing polyacrylamide gels after separation by micro-disc-electrophoresis different deoxyribonucleases are detectable in bull seminal plasma. There are two groups of acid deoxyribonuclease-activities with a pH optimum at pH 5.0, one with a pH optimum at pH 7.4 and an additional one with a pH optimum at pH 8.5.

Characterization of different deoxyribonucleases in human lymphocytes.

Four groups of deoxyribonuclease activities from human lymphocytes have been characterized by deoxyribonuclease assay in DNA-containing polyacrylamide gels following their separation by disc-electrophoresis. All activities hydrolyse DNA endonucleolytically. One neutral deoxyribonuclease found in the cytoplasmic fraction prefers native or UV-irradiated DNA over denatured DNA as substrate and is a 5'-monoester former. Two groups of acid deoxyribonuclease activities are detectable in the nuclear fraction. Both are 3'-monoester formers. One is as well active with denatured DNA as with native DNA, the other one shows the same activity with native and UV-irradiated DNA but lower activity with denatured DNA. An alkaline deoxyribonuclease activity, also localized in the nucleus, is a 5'-monoester former, and prefers denatured or UV-irradiated DNA as substrate.

Deoxyribonuclease activities in human leukemic cells and mouse lymphoblasts

The deoxyribonuclease activities from human lymphocytes have been compared with the activities from acute lymphocytic leukemic cells and mouse leukemic cells L5178Y using the in situ detection of deoxyribonucleases in DNA-containing polyacrylamide gels following their separation by micro-disc-electrophoresis. A neutral deoxyribonuclease activity is completely missing in leukemic cells of untreated patients while a group of acid deoxyribonuclease activities is increased. A similar deoxyribonuclease pattern can be seen in L5178Y cells. Under medical treatment the increment of the acid deoxyribonuclease activities disappears and the neutral deoxyribonuclease activity reappears.

Effect of Exogenous DNA on Acid Deoxyribonuclease Activity in Intact Roots of Vicia faba L.

Effect of Exogenous DNA on Acid Deoxyribonuclease Activity in Intact Roots of Vicia faba L.

A comparison of the biochemical changes induced in mouse brain by cuprizone toxicity and by scrapie infection

Studies were made of the activity of acid deoxyribonuclease and eleven glycoside hydrolases, and the rate of incorporation of DNA and glycoprotein precursors in the brains of mice fed on a diet containing 0·5 per cent. cuprizone (biscyclohexanoneoxaldihydrazone) for up to 16 weeks. The profile of changes observed was very similar to that found in the brains of mice infected with the Chandler strain of scrapie agent. The biochemical lesions of cuprizone toxicity in brain could be reversed by withdrawing cuprizone from the diet. It is concluded that many of the biochemical changes seen in Chandler scrapie are not specific for the disease and that cuprizone toxicity may be a useful control for certain types of scrapie experiment.

Fibroma virus-induced acid deoxyribonuclease in rabbit kidney cells.

Acid deoxyribonuclease (DNase) activity increased 6 to 10 fold in primary rabbit kidney (PRK) cells infected with Shope rabbit fibroma virus. Increase in enzyme synthesis was proportional to viral DNA replication. Addition of actinomycin D or puromycin to cultures terminated enzyme synthesis, indicating requirement for messenger RNA andde novo protein synthesis. The enzyme which was concentrated by ammonium sulfate precipitation and purified by DEAE cellulose chromatography degraded single and double-stranded DNA in a ratio of 2∶1 and required Mg++ ions for optimal activity. It was inactivated by more than 80 per cent in 5 minutes at 50° C and the presence of deoxynucleoside triphosphates in a reaction mixture inhibited its activity.

Endocytosis and the adaptive acid hydrolases in Tetrahymena pyriformis GL.

Endocytosis of yeast cells by Tetrahymena pyriformis GL for a period of 2.5 h produced changes in cellular acid hydrolases. Acid phosphatase, acid deoxyribonuclease and acid proteinase activities were markedly increased, whereas there was a decrease in acid ribonuclease activity and little change in α-glucosidase activity. These alterations do not appear to be due to any alteration in the rates of secretion of these enzymes into the milieu. Evidence is presented that the cellular enzyme increases found upon endocytosis of yeast reflect changes in lysosomal enzymes, because it was shown that the acid phosphatase activity increase resulted in an increased amount of latent enzyme within the cell. The results also support the idea that there are at least 3 distinct populations of lysosomes, in addition to phagolysosomes, present in Tetrahymena pyriformis GL, with different modes of formation. There appears to be a large excess of lysosomes, uncombined with phagosomes, present in these fed cells since latency averaged 66% in broken-cell preparations which contained very few intact phagolysosomes. The phagolysosomal acid phophatase activity cannot account for more than 34% of that present in the cell. The endocytosis of yeast in the presence of growth medium resulted in a marked drop in the rate of cell division as compared to cells growing in the growth medium alone. The results are discussed.

Distribution of lysosomal enzymes between parenchymal and Kupffer cells of rat liver

1. 1. Suspensions of rat liver cells were prepared by perfusing the liver in vitro with solutions containing collagenase and hyaluronidase. About 90% of the liver mass was recovered in the cell suspension. 2. 2. Parenchymal cells were prepared by differential centrifugation or by magnetic removal of iron-loaded Kupffer cells. About 60% of the liver mass was regained in the parenchymal fractions. Specific activity of β glucoronidase (EC 3.2.1.31) was about the same in the parenchymal cells and in liver homogenates. Specific activity of acid deoxyribonuclease (EC 3.1.4.6) in parenchymal cells was about 65% of that in liver homogenates. 3. 3. Kupffer cells were separated from parenchymal cells either by incubating the original cell suspension with pronase, which destroys the parenchymal cells, or by magnetic means. 70 and 40% of the Kupffer cells in the original cell suspension could be recovered by the pronase method and the magnet method, respectively. Iron-loaded cells were contaminated with parenchymal cells. Acid deoxyribonuclease and β-glucuronidase were found to be selectively concentrated in Kupffer cells prepared with pronase. Specific activities of β-glucuronidase and acid deoxyribonuclease were, respectively, 3.6 and 13.6 times higher in pronase-prepared Kupffer cells than in parenchymal cells. Only acid deoxyribonuclease was found to be increased in Kupffer cells isolated by means of a magnet (about twice as high as in parenchymal cells).