Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.4.37 (CNPase)
 539 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of postmortem autolysis in situ on myelin proteins and glycoproteins were studied in 25- and 125-day-old mouse brain and in adult bovine brainstem. In bovine myelin a loss of the major myelin glycoprotein was the only difference observed when the tissue was left at 19 degrees C for 24 hours compared to immediately frozen material. In the autolysed mouse brain, the myelin major glycoprotein was the most affected component with a 55% decrease. Both myelin basic protein components were degraded with a 35% loss. The other myelin proteins did not change under the conditions used for this study. There was also no change in the specific activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase, a myelin-associated enzyme. Using the double labelling technique with [3H]fucose and [3 5S] sulfate as precursors injected intracranially, a shift of the major myelin glycoprotein labelled with radioactive sulfate towards a smaller apparent molecular size was observed as a result of the autolysis whereas the electrophoretic mobility of the fucose labelled major peak was unaffected.
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PMID:Changes in CNS myelin proteins and glycoproteins after in situ autolysis. 19 16

Myelin purified from the central nervous system of Xenopus laevis contained the same major lipid and protein components as human myelin. However, some minor differences in the myelin proteins were noted. The Xenopus basic protein had a higher apparent mol wt. on sodium dodecyl sulfate gels than the corresponding mammalian protein. The absolute specific activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase in the Xenopus myelin was considerably higher than in mammals. There were differences in the high mol wt. proteins, and the glycoproteins in Xenopus myelin were more heterogeneous than those in mammals. Peripheral myelin from Xenopus sciatic nerve was compared with that from the rat. The lipids in the two types of myelin were similar. There was a major glycoprotein in the Xenopus myelin corresponding to the P0 protein and a basic protein of slightly larger mol wt. than the P1 protein of rat myelin.
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PMID:A biochemical comparison of Xenopus laevis and mammalian myelin from the central and peripheral nervous systems. 21 Dec 3

The brains from 1-week-old rat pups were used to prepare cultures of glial cells. After 24 hr in culture the cells were changed to a chemically defined serum-free medium (CDM). We have used antibodies against gangliosides (monoclonals A2B5 and LB1) and against galactocerebrosides to monitor the influence of insulin on the development of oligodendrocytes from precursor cells. In these oligodendrocyte-enriched cultures we investigated the influence of insulin and IGF-I on the activity of the marker enzymes 2',3'-cyclic-nucleotide 3'-phosphodiesterase (CNPase) and glycerol-3-phosphate dehydrogenase (G3PDH), and on lipid metabolism. 1) Incorporation of [35S] sulfate into sulfolipids was stimulated by insulin (optimal concentration 100 nM). A sharp peak in sulfolipid synthesis was seen at day 5-6 in culture. 2) Insulin stimulated the expression of CNPase and G3PDH in culture. 3) The stimulating effect of insulin on sulfolipid synthesis, CNPase, and G3PDH activity was mimicked by IGF-I (13 nM). 4) The incorporation of [35S] into sulfolipids and [2-3H]acetate into fatty acids and cholesterol was reversibly reduced by temporary omission of insulin from the medium. These results indicate that insulin and IGF-I stimulate the development of oligodendrocytes in our culture system and that insulin has a general anabolic effect on the lipid metabolism of the cells.
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PMID:Effects of insulin and insulin-like growth factor (IGF-I) on oligodendrocyte-enriched glial cultures. 283 44

Oligodendrocytes generate myelin as extensions of the plasma membrane. Myelin has been well characterized, yet little is known concerning oligodendrocyte plasma membrane. We have developed a reproducible method for the isolation of an oligodendrocyte plasma membrane-rich fraction (F2.2). F2.2 has a 25-fold enrichment in K+-dependent p-nitrophenyl phosphatase, a plasma membrane marker. Impurities are composed of Golgi elements (8-12%), microsomes (4-6%), and lysosomal membranes (1-5%). Our starting material was oligodendrocytes kept in culture in a nonattached state for 3 to 5 d. After disrupting the cells and removing nuclei (P1), the supernatant (SP1) was fractionated on a self-generating 20% Percoll gradient into three bands: F1, F2, and F3. F1 had only 3% of the applied protein and was not characterized. F2, with 11% of the protein, was fivefold enriched in plasma membrane. F3 had 27% of initial protein; it consisted of a crude mitochondrial and lysosomal fraction. F2 was further purified by first washing it hypotonically, treating it with Mg2+, and then fractionating it on a Ficoll step gradient that yielded F2.2 at the interphase. Morphologically F2.2 comprises 1) membranous sheets, often with more than one membrane in close apposition; 2) membrane vesicles of various sizes and shapes frequently filled with amorphous material; 3) Golgi elements; and 4) unrecognizable profiles. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profile of F2.2 reveals CNPase as a major component in agreement with the high CNPase specific activity (3,860 mumol/mgP/h) found in F2.2. Other significant polypeptides have Mr = 170,000, 135,000, 108,000, 80,000, 53,000, 38,500, 32,000, and 22,200.
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PMID:Plasma membrane of cultured oligodendrocytes: I. Isolation, purification, and initial characterization. 297 38

Monoclonal antibody against 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNP) was generated by fusing mouse myeloma cells with spleen cells from BALB/c mice immunized with delipidated white matter from rat corpus callosum. The antibody was characterized by solid-phase radioimmunoassay, immunoblot of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), immunoprecipitation from C6 glioma cells, and indirect immunofluorescence staining of monolayer cultures containing oligodendrocytes. The monoclonal antibody bound specifically to an intracellular antigen of oligodendrocytes, but not to Schwann cells, astrocytes, neurons, or fibroblast cytoplasm. The immunoblot of SDS-PAGE of CNS myelin showed that the antibody identified two protein bands at 48,000 and 50,000 molecular weight. These proteins were not identified in peripheral nervous system myelin. The monoclonal antibody immunoprecipitated CNP enzyme activity from extracts of C6 glioma cells. This monoclonal antibody should prove useful in further study of this myelin-specific enzyme in CNS myelin and in cells responsible for myelin production.
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PMID:A monoclonal antibody raised to corpus callosum extract reacts with 2',3'-cyclic nucleotide 3'-phosphohydrolase. 299 39

Biochemical and morphological studies of myelin subfractions were undertaken on Lewis rats during the early stage of the development of experimental allergic encephalomyelitis (EAE). Myelin subfractions, obtained by sucrose density gradient centrifugation at 10 days post-induction, were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and assayed for 2':3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) activity. Aliquots were processed for electron microscopic analysis. When comparing the myelin subfractions of EAE-affected animals with those of controls, differences were observed only in the light fractions, i.e., a decrease in the specific activity of CNPase and in the percentage of basic proteins relative to the total proteins of the fraction. This decrease was also evident in the basic protein/proteolipid protein ratio which is frequently used in the literature. In addition, electron microscopic observations demonstrated strong differences in the morphology of the same fraction. These findings suggest that the light fraction is the most sensitive in the early stages of the disease and must play a key role in demyelinating processes.
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PMID:Biochemical changes in central nervous system membranes in experimental allergic encephalomyelitis. 301 92

Cultures of glial cells were prepared from the brains of one-week-old rat pups. After one day in culture, serum was omitted from the medium and replaced by a combination of growth-stimulating hormones and other factors that enhanced the percentage of oligodendrocytes in the cultures. We investigated the effects of hydrocortisone on the development of oligodendrocytes, on the activities of oligodendrocyte-specific enzymes and on glucose- and lipid-metabolism of the glial cells. Hydrocortisone greatly enhanced the survival of glial cells in culture. The development of galactocerebroside-positive cells and the specific activity of 2',3'-cyclic-nucleotide 3'-phosphodiesterase were stimulated by 50 nM hydrocortisone, whereas these effects were partly reversed at higher concentrations of the hormone. The specific activity of glycerol-3-phosphate dehydrogenase was markedly stimulated by hydrocortisone; 1 microM or higher concentrations of hydrocortisone were required for an optimal effect. The consumption of glucose and the production of lactate were lowered by hydrocortisone whereas the oxidation of [6-14C]glucose to 14CO2 was not affected. Incorporation of [35S]sulfate into sulfolipids was greatly enhanced by hydrocortisone and [14C]incorporation from [1-14C]acetate into cholesterol and fatty acids was also stimulated but to a smaller extent. These results show that hydrocortisone exerts a general trophic function on glial cells in our culture system; enhances the ratio of oligodendrocytes over astrocytes, possibly by directing bipotential progenitor cells to develop into oligodendrocytes; specifically induces glycerol-3-phosphate dehydrogenase in oligodendrocytes.
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PMID:Hydrocortisone stimulates the development of oligodendrocytes in primary glial cultures and affects glucose metabolism and lipid synthesis in these cultures. 304 Jan 87

Heterosis (hybrid vigor) for brain myelin content has been examined in detail in (C57BL/6J x DBA/2J)F1 hybrid mice at 17 days of age. The amount of myelin isolated from the F1 hybrid brain is greater than that isolated from either parental strain. In addition, the total protein content in the myelin of the three genotypes showed the following trend: F1 greater than DBA greater than C57. However, no discernible differences in myelin protein compositions could be detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Analysis of the whole brain for several myelin-associated constituents such as GM1 ganglioside, 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNPase), 5'-nucleotidase, and carbonic anhydrase indicated that heterosis exists for these components. No heterosis was found for such nonmyelin constituents as gangliosides GD1a, GT, GQ, RNA, DNA, and choline acetyltransferase. A developmental study of the whole brain CNPase indicated that the heterotic effect was greatest during the most active period of myelination (17-30 days). We conclude that the heterotic effect is specific for myelin content and is probably the result of an accelerated myelin synthesis. The heterotic effect should have great potential as a new model for studying aspects of myelinogenesis.
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PMID:Biochemical study of heterosis for brain myelin content in mice. 618 36

Myelin was purified from the spinal cords of normal mice and mice heterozygous for the shiverer mutation, and measurements were made of the major myelin proteins and lipids and the specific activities of three myelin-associated enzymes. The myelin purified from the spinal cords of the heterozygotes (shi/+) was deficient by 30-40% in yield and had an apparently unique composition. In particular, when compared with normal mouse spinal cord myelin, there were more high-molecular-weight protein, less myelin basic protein, a higher protein-to-lipid ratio, and higher specific activities of 2',3'-cyclic nucleotide-3'-phosphohydrolase (EC 3.1.4.37) and carbonic anhydrase (EC 4.2.1.1) in the myelin purified from the shi/+ animals. These abnormalities were reflected in the composition of shi/+ whole spinal cord, where the protein-to-lipid ratio was intermediate between the respective values for +/+ and shi/shi spinal cords. Whole brains from shi/+ mice showed deficiencies in galactocerebroside and galactocerebroside sulfate and an increase in total phospholipid, and the lipid composition in the brains of the shi/shi mice was similar to that reported for another dysmyelinating mutant, quaking. The findings provide the first values for the lipids in normal mouse spinal cord myelin and show that heterozygotes are affected by the shiverer mutation. The observations imply that there can be considerable deviation from the normal CNS myelin content and composition without apparent qualitative morphological abnormalities or loss of function and that the amount of myelin basic protein available during myelination may influence the incorporation of other constituents into the myelin membranes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Biochemical abnormalities in spinal cord myelin and CNS homogenates in heterozygotes affected by the shiverer mutation. 620 May 71

Synaptosomes were prepared from rat cortex by subjecting a washed crude mitochondrial pellet to centrifugation first on discontinuous Ficoll-isotonic sucrose gradients and then on discontinuous sucrose gradients. The synaptosome fraction, collected from the 7.5-14% Ficoll band (II), was further separated into two additional fractions, designated IIA and IIB, which bank at the 0.32-1.05 M and at the 1.05-1.6 M sucrose interfaces, respectively. Electron microscopic analysis showed that fraction IIB contained synaptosomes and extra terminal mitochondria and was essentially free of membrane fragments. Further characterization showed that IIB contained 69% of the protein and 83% of the lactic dehydrogenase activity of fraction II and had a specific activity of a 2',3'-cyclic nucleotide 3'-phosphohydrolase approximately 1% of that obtained with myelin. Fraction IIA had approximately 50% the specific activity of the 2',3'-cyclic nucleotide 3'-phosphohydrolase found in myelin. Synaptic plasma membranes were prepared by lysing fraction IIB in 1 mM sodium phosphate, 0.1 mM EDTA at pH 8.5 and subjecting this preparation to centrifugation on a discontinuous sucrose density gradient. Enzymatic analysis indicated that membranes banding at the 0.6-0.8 M sucrose interface had high specific activities of plasma membrane enzymes (e.g. acetylcholinesterase, ATPase, 5'-nucleotidase). The specific activity of the (Na+ + K+)-ATPase in the purified membrane preparation was 8-fold higher than that in the original homogenate. Specific activities of various marker enzymes indicated that the composition of these membrane preparations for the most part was synaptic plasma membranes, approximately 7% mitochondrial outer membranes and 3% a membrane containing 2',3'-cyclic nucleotide 3'-phosphohydrolase activity. The polypeptide compositions of three possible contaminating membranes and of synaptic membranes were compared by electrophoresis in 6-20% gradient polyacrylamide gels in the presence of sodium dodecyl sulfate. Whereas mitochondrial and myelin membranes had distinct compositions, the compositions of the microsomal and synaptosomal plasma membranes were similar. Synaptic plasma membranes contained at least 27 polypeptides; the three major polypeptides had molecular weights of 103,000; 54,000; and 50,000. The major polypeptides of soluble synaptosomal proteins had molecular weights of 54,000 and 42,000.
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PMID:An improved method of preparing rat brain synaptic membranes. Elimination of a contaminating membrane containing 2',3'-cyclic nucleotide 3'-phosphohydrolase activity. 624 53


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