共查询到20条相似文献,搜索用时 15 毫秒
1.
Purified rat brain Ca2+/calmodulin-dependent protein kinase II (CaM-kinase II) is stimulated by brain gangliosides to a level of about 30% the activity obtained in the presence of Ca2+/calmodulin (CaM). Of the various gangliosides tested, GT1b was the most potent, giving half-maximal activation at 25 microM. Gangliosides GD1a and GM1 also gave activation, but asialo-GM1 was without effect. Activation was rapid and did not require calcium. The same gangliosides also stimulated the autophosphorylation of CaM-kinase II on serine residues, but did not produce the Ca2+-independent form of the kinase. Ganglioside stimulation of CaM-kinase II was also present in rat brain synaptic membrane fractions. Higher concentrations (125-250 microM) of GT1b, GD1a, and GM1 also inhibited CaM-kinase II activity. This inhibition appears to be substrate-directed, as the extent of inhibition is very dependent on the substrate used. The molecular mechanism of the stimulatory effect of gangliosides was further investigated using a synthetic peptide (CaMK 281-309), which contains the CaM-binding, inhibitory, and autophosphorylation domains of CaM-kinase II. Using purified brain CaM-kinase II in which these regulatory domains were removed by limited proteolysis. CaMK 281-309 strongly inhibited kinase activity (IC50 = 0.2 microM). GT1b completely reversed this inhibition, but did not stimulate phosphorylation of the peptide on threonine-286. These results demonstrate that GT1b can partially mimic the effects of Ca2+/CaM on native CaM-kinase II and on peptide CaMK 281-309. 相似文献
2.
James R. Goldenring Laura C. Otis Robert K. Yu Robert J. DeLorenzo 《Journal of neurochemistry》1985,44(4):1229-1234
The effects of gangliosides on phosphorylation were studied in rat brain membrane. Gangliosides stimulated phosphorylation only in the presence of Ca2+ with major phosphoproteins of 45,000, 50,000, 60,000, and 80,000 daltons and high-molecular-weight species. In addition, gangliosides inhibited the phosphorylation of three proteins with molecular weights of 15,000, 20,000, and 78,000 daltons. The two low-molecular-weight proteins comigrated with rat myelin basic proteins. Ganglioside stimulation was dependent on the formation of a Ca2+-ganglioside complex since the calcium salt of gangliosides stimulated phosphorylation maximally. Disialo and trisialo gangliosides were more potent stimulators of kinase activity than the monosialo GM1 X GD1a was the most potent activator tested. Asialo-GM1, cerebroside, sialic acid, neuraminyllactose, sulfatide, and the acidic phospholipids phosphatidylserine and phosphatidylinositol did not stimulate kinase activity. The Ca2+-dependent, ganglioside-stimulated phosphorylation was qualitatively similar to the pattern for calmodulin-dependent phosphorylation. However, while calmodulin-dependent kinase activity was inhibited with an IC50 of 10 microM trifluoperazine, ganglioside-stimulated kinase was inhibited with an IC50 of 200 microM trifluoperazine. These results indicate that gangliosides have complex effects on membrane-associated kinase activities and suggest that Ca2+-ganglioside complexes are potent stimulators of membrane kinase activity. 相似文献
3.
Activation of protein kinase C by ganglioside GM3 in the presence of calcium and 12-O-tetradecanoylphorbol-13-acetate 总被引:1,自引:0,他引:1
T Momoi 《Biochemical and biophysical research communications》1986,138(2):865-871
Ganglioside GM3 and 12-O-tetradecanoylphorbol-13-acetate activated protein kinase C as substitutes for phosphatidylserine and diacylglycerol. Hydrophobic gangliosides such as GM4 and GM3 were rather more potent activators of protein kinase C than hydrophilic ones such as GD1a and GT1b. Active tumor promoters such as teleocidin, mezerein, phorbol 12,13-acetate and phorbol 12,13-dibenzoate also activated protein kinase C, but not inactive tumor promoters such as phorbol and 4-alpha-phorbol 12,13-didecanoate. 相似文献
4.
A ganglioside-stimulated protein phosphorylation system was discovered in plasma membrane fractions of human neuroblastoma cells (GOTO). Gangliosides (GQ1b, GT1a, GT1b, GD1a, GD1b, GD3, and GM1) could stimulate this system. GQ1b showed the most effective stimulation among these gangliosides. The substrate specificity was rather broad. Not only some (de novo) proteins of the membranes but also purified histones and tubulin were phosphate-acceptable. This protein phosphorylation system specifically depended upon Ca2+ (optimum concentration: 50-100 microM). The optimum pH was 7.0-7.5. GQ1b/Ca2+ could not directly activate well known protein kinases (Ca2+/phospholipid-activated protein kinase, Ca2+/calmodulin-activated protein kinase, and cyclic nucleotide-dependent protein kinases). Furthermore, GQ1b could replace neither phospholipids nor calmodulin. Thus, an unknown, new type of protein kinase(s) may be involved in this system. Alternatively, GQ1b may activate some known protein kinase(s) in cooperation with another unknown factor which may be removed during the preparation of the partially purified known protein kinase used in this experiment. 相似文献
5.
Ganglioside-modulated protein phosphorylation. Partial purification and characterization of a ganglioside-stimulated protein kinase in brain 总被引:7,自引:0,他引:7
K F Chan 《The Journal of biological chemistry》1987,262(11):5248-5255
Gangliosides have profound modulatory effects on protein phosphorylation in brain. A protein kinase activated directly by gangliosides has been partially purified from the particulate fractions of guinea pig brain through extraction with nonionic detergent, ion-exchange chromatography, hydrophobic chromatography, and gel filtration. This novel ganglioside-stimulated protein kinase is distinct from cAMP-dependent, Ca2+/calmodulin-dependent, and Ca2+/phospholipid-dependent protein kinases. The partially purified kinase preparation could undergo ganglioside-stimulated autophosphorylation of a major phosphoprotein with Mr corresponding to 68,000. It also could phosphorylate exogenous substrates such as the synthetic peptide Leu-Arg-Arg-Ala Ser-Leu-Gly. The requirement of gangliosides for the activation of kinase activity is dose-dependent and specific. Among the various gangliosides tested, GT1b and GD1a were found to be the most potent activators, whereas GD1b and GM1 were slightly less effective. The activation process is rapid and does not require the presence of Ca2+, suggesting that the stimulatory effect of gangliosides is not mediated through limited proteolysis or Ca2+-glycolipid complexes. Although the exact physiological significance of the ganglioside-stimulated protein kinase is not known at present, it is possible that certain functions related to gangliosides in the nervous system are mediated through the activation of this novel enzyme. 相似文献
6.
Myelin-associated glycoprotein (MAG, Siglec-4) is a quantitatively minor membrane component expressed preferentially on the innermost myelin wrap, adjacent to the axon. It stabilizes myelin-axon interactions by binding to complementary ligands on the axolemma. MAG, a member of the Siglec family of sialic acid-binding lectins, binds specifically to gangliosides GD1a and GT1b, which are the major sialoglycoconjugates on mammalian axons. Mice with a disrupted Galgt1 gene lack UDP-GalNAc:GM3/GD3 N-acetylgalactosaminyltransferase (GM2/GD2 synthase) and fail to express complex brain gangliosides, including GD1a and GT1b, instead expressing a comparable amount of the simpler gangliosides GM3, GD3, and O-acetyl-GD3. Galgt1-null mice produce similar amounts of total myelin compared to wild-type mice, but as the mice age, they exhibit axon degeneration and dysmyelination with accompanying motor behavioral deficits. Here we report that Galgt1-null mice display progressive and selective loss of MAG from the brain. At 1.5 months of age, MAG expression was similar in Galgt1-null and wild-type mice. However, by 6 months of age MAG was decreased approximately 60% and at 12 months of age approximately 70% in Galgt1-null mice compared to wild-type littermates. Expression of the major myelin proteins (myelin basic protein and proteolipid protein) was not reduced in Galgt1-null mice compared to wild type. MAG mRNA expression was the same in 12-month-old Galgt1-null compared to wild-type mice, an age at which MAG protein expression was markedly reduced. We conclude that the maintenance of MAG protein levels depends on the presence of complex gangliosides, perhaps due to enhanced stability when MAG on myelin binds to its complementary ligands, GD1a and GT1b, on the apposing axon surface. 相似文献
7.
Arun K. Chakrabarti Somsankar Dasgupta Naren L. Banik Edward L. Hogan 《Journal of neurochemistry》1990,54(5):1816-1819
We examined ganglioside modulation of the activity of the millimolar Ca2(+)-sensitive form (mCANP) of calcium-activated neutral proteinase (CANP), which is enriched in myelin, from brain. GM1, GD1a, GT1a, GM2, and GM4 produced a concentration-dependent increase of mCANP activity. GD1a stimulated the greatest increase of enzyme activity (107%), followed by GT1a, whereas GD1b was inhibitory (56%). GM1, GM2, and GM4 stimulated but less so than GD1a and GT1a. Free N-acetylneuraminic acid, asialo-GM1, GM3, and a ganglioside mixture containing GM1, GD3, GD1a, and GD1b had no effect. The ganglioside-mediated modulation was not affected by trifluoperazine and chlorpromazine (phospholipid-binding antagonists). The mCANP Ca2+ requirement was significantly reduced in the presence of stimulatory gangliosides, and this increased sensitivity varied (10-50-fold) with ganglioside structure. Gangliosides may interact with membrane mCANP and modulate its proteolytic action. 相似文献
8.
We systematically examined the effects of gangliosides on the plasma membrane Ca(2+)-ATPase (PMCA) from porcine brain synaptosomes. Our results showed that GD1b (two sialic acid residues) stimulated the activity, GM1 (one sialic acid residue) slightly reduced the activity, while asialo-GM1 (no sialic acid residue) markedly inhibited it, suggesting that sialic acid residues of gangliosides are important in the modulation of the PMCA. We also examined the oligosaccharide effects by using GM1, GM2, and GM3 whose only difference was in the length of their oligosaccharide chain. GM1, GM2, and GM3 reduced the enzyme activities, whereas GM2 and GM3 were potent inhibitors. Gangliosides affect both affinity for Ca(2+) and the Vmax of enzyme. It was observed that GD1b and GM2 increased the affinity of the enzyme for Ca(2+). GD1b, GM2 affected the Vmax with an increase of GD1b, but decreases of GM2. The study of the affinity for ATP and the Vmax of enzyme in the presence of gangliosides showed that GD1b and GM2 had little effect on the ATP binding to the enzyme, but the Vmax was apparently changed. Moreover, the effects of gangliosides are additive to that of calmodulin, suggesting that the modulation of PMCA by gangliosides should be through a different mechanism. The conformational changes induced by gangliosides were probed by fluorescence quenching. We found that fluorescent quenchers (I(-) and Cs(+)) with opposite charges had different accessibility to the IAEDANS binding to the PMCA in the presence of gangliosides. An apparent red shift (25nm) with increased maximum of fluorescence spectrum was also observed in the presence of GD1b. 相似文献
9.
Paola Palestini Massimo Masserini Amelia Fiorilli Emanuela Calappi Guido Tettamanti 《Journal of neurochemistry》1993,61(3):955-960
Abstract: Age-related changes of the ceramide composition of gangliosides were studied in the synaptosomal and myelin fractions from rat brain, carrying plasma membranes of neuronal and glial origin, respectively. The five major gangliosides (GM1, GD1 a, GD1 b, GT1 b, and GQ1 b) present in these fractions were separated and quantitated by normal-phase HPLC. Each ganglioside was then fractionated by reverse-phase HPLC into the molecular species carrying a single long-chain base (LCB). The largely preponderant LCBs in the synaptosomal and myelin fractions were the C18:1 and C20:1. The content of C20.1 LCB, generally low at 1 month, increased with age in all analyzed gangliosides and in all subcellular fractions and was greater in the "b series" than in the "a series" gangliosides. Remarkably, GM1 was the only ganglioside where the proportion of LCB 20:1 was higher in the synaptosomal fraction than in the myelin fraction. The fatty acid composition of the C18:1 or C20:1 LCB species of the different gangliosides in the synaptosomal and myelin fractions did not undergo appreciable changes with age. Stearic acid was largely predominant in all the gangliosides of the synaptosomal fraction, more in the C18:1 than in the C20:1 LCB species (80–90% vs. 60–70%). The gangliosides of the myelin fraction were characterized by a lower content of 18:0 and a much higher content of 16:0 and 18:1 fatty acids than those of the synaptosomal fraction. Thus, the ceramide composition is different in the gangliosides of neuronal and myelin origin and appears to be subjected to an age-related control. 相似文献
10.
Regulation of protein kinase C by lysophospholipids. Potential role in signal transduction 总被引:13,自引:0,他引:13
Certain lysophospholipids, lysophosphatidylcholine (lyso-PC) in particular, stimulated protein kinase C at low concentrations (less than 20 microM) but, conversely, inhibited it at high concentrations (greater than 30 microM). Protein kinase C stimulation by lyso-PC required the presence of phosphatidylserine (PS) and Ca2+ and was associated with a decreased Ka for PS and increased Ka for Ca2+ of the enzyme. Cardiolipin and phosphatidic acid could partially substitute for PS in supporting the stimulatory effect of lyso-PC. Lyso-PC also biphasically regulated protein kinase C activated by diolein. Of several synthetic lyso-PC preparations tested, the oleoyl, myristoyl and palmitoyl derivatives were most active. Data from the Triton X-100 mixed micellar assay indicated that 1.4 and 14.0 mol of lyso-PC/micelle produced a maximal stimulation and a complete abolishment of the stimulation of protein kinase C, respectively. Protein kinase C stimulation by lyso-PC, with a pH optimum of about 7.5, was observed for phosphorylation of histone H1, myelin basic protein, and the 35- and 47-kDa proteins from the rat brain, but not for that of other histone subfractions and protamine. Lyso-PC acted synergistically with diacylglycerol in stimulating protein kinase C, whereas the stimulation by lyso-PC was additive to that by oleic acid. Protein kinase C inhibitors (alkyllysophospholipid, sphingosine, tamoxifen, and polymyxin B) inhibited more potently the protein kinase C activity stimulated by PS/Ca2+/lyso-PC than that stimulated by PS/Ca2+. The stimulatory and inhibitory effects of lyso-PC were not observed for myosin light chain kinase and cAMP-dependent protein kinase, indicating a specificity of its actions. The present findings suggested that lyso-PC, likely derived from membrane PC by the action of phospholipase A2, might play a role in signal transduction via a dual regulation of protein kinase C, and that it could further modulate the enzyme and hence the cellular activity by interplaying with diacylglycerol and unsaturated fatty acid, the two other classes of cellular mediators also shown to be activators of protein kinase C. 相似文献
11.
Specific immune damage to liposomes containing Forssman or globoside glycolipid was inhibited when the liposomes also contained ganglioside. The activity of a human monoclonal Waldenstr?m macroglobulin antibody to Forssman glycolipid was inhibited by each of three gangliosides tested, GM3, GD1a and GD1b. Inhibition of the monoclonal antibody was dependent on the amount of ganglioside in the liposomes, and was diminished by reducing the relative amount of ganglioside. Inhibition also correlated positively with the number of ganglioside sialic acid groups, with inhibition by GT1b greater than GD1a greater than GM3. Naturally occurring human antibodies to globoside glycolipid were detected in 18% (9 out of 50) of normal human sera tested. Immune damage to liposomes induced by each of the three highest-reacting human anti-globoside sera was blocked by liposomal GM3. We conclude that gangliosides can strongly influence immune damage to membranes induced by antibody interactions with adjacent neutral glycolipids. 相似文献
12.
Abstract: Phosphorylation of the nervous system-specific protein GAP-43 in growth cones in vivo increases as the growth cones near their targets, at a time when the gangliosides GM1 and GD1a are being accumulated in the growth cone membrane, thus raising the possibility that the gangliosides could modulate GAP-43 behavior. We used a subcellular fraction of intact isolated growth cones to show that both GM1 and GD1a affected the calcium- dependent posttranslational regulation of GAP-43 in several similar ways. Both gangliosides induced rapid incorporation of phosphate into GAP-43; however, the induction was undetectable with our antibody 2G12 that is specific for kinase C-phosphorylated GAP-43. Furthermore, neither ganglioside stimulated kinase C activity in isolated growth cones, suggesting that the rapid Phosphorylation may not be on Ser41 , the kinase C site. However, both gangliosides did induce a slower accumulation of GAP-43 phosphorylated on Ser41 , apparently by inhibiting a phosphatase. Finally, calcium-dependent proteolysis of GAP-43 was also stimulated by both GM1 and GD1a. In contrast, GD1a, but not GM1, caused the redistribution of GAP-43 into the isolated growth cone cytoskeleton. The results demonstrate that both gangliosides can modulate the calcium-dependent regulation of GAP-43. 相似文献
13.
Gangliosides are known to be differentiation-inducing molecules in mammalian stem cells. We studied the interaction between
the molecular structure of glycosphingolipids (GSLs) and their promoting mechanisms of the phagocytic processes in human polymorphonuclear
leukocytes (PMN). The effect of various gangliosides from mammalian tissues on adhesion, phagocytosis, phagosome–lysosome
(P–L) fusion and superoxide anion production was examined by human PMN using heat-killed cells of Staphylococcus aureus coated
with GSLs. Gangliosides GM3, GD1a, GD3 and GT1b showed a marked stimulatory effect on the phagocytosis and P–L fusion in a
dose-dependent manner, while ganglioside GM1, asialo GM1 and neutral GSLs did not. The relative phagocytic rate of ganglioside
GM3-coated S. aureus was the highest among the tested GSLs. Both P–L fusion rate and phagocytosis of S. aureus were elevated
significantly when coated with ganglioside GD1a, GD3 or GT1b, and GT1b gave a five times higher rate than that of the non-coated
control. These results suggest that the terminal sialic acid moiety is essential for the enhancement of phagocytosis and that
the number of sialic acid molecules in the ganglioside is related to the enhancement of the P–L fusion process. On the other
hand, the superoxide anion release from PMN was not affected by ganglioside GM2, GM3, GD1a or GT1b. Furthermore, to clarify
the trigger or the signal transduction mechanism of phagocytic processes, we examined the effect of protein kinase inhibitors
such as H-7, staurosporine (protein kinase C inhibitor), H-89 (protein kinase A inhibitor), genistein (tyrosine kinase inhibitor),
ML-7 (myosin light chain kinase inhibitor), and KN-62 (Ca2+/calmodulin-dependent protein kinase II inhibitor) on ganglioside-induced
phagocytosis. H-7, staurosporine and KN-62 inhibited ganglioside-induced phagocytosis in the range of concentration without
cell damage, while H-89, genistein and ML-7 did not. Moreover, H-7 and KN-62 inhibited ganglioside-induced P–L fusion. These
results suggest that protein kinase C and Ca2+/calmodulin-dependent protein kinase II may be involved in the induction of
phagocytosis and P–L fusion stimulated by gangliosides.
This revised version was published online in November 2006 with corrections to the Cover Date. 相似文献
14.
Among calmodulin-non-binding glycosphingolipids, GM3, sialosylneolactotetraosylceramide (LM1), and sulfatide potently activated calmodulin-dependent cyclic nucleotide phosphodiesterase with or without Ca2+ showing ED50 1-5 microM. In contrast to calmodulin-binding gangliosides, these glycosphingolipids activated the enzyme up to the maximum level achieved by Ca2+/calmodulin and did not inhibit the activity at higher concentrations. Competition studies with GD1b that bind both to calmodulin and the enzyme suggest that the calmodulin-non-binding glycosphingolipids activate the enzyme through interaction with the same site of the enzyme as GD1b interacts. 相似文献
15.
Erik A Miljan Emmanuelle J Meuillet Barbara Mania-Farnell David George Hirotaka Yamamoto Hans-Georg Simon Eric G Bremer 《The Journal of biological chemistry》2002,277(12):10108-10113
Ganglioside GM3 inhibits epidermal growth factor (EGF)-dependent cell proliferation in a variety of cell lines. Both in vitro and in vivo, this glycosphingolipid inhibits the kinase activity of the EGF receptor (EGFR). Furthermore, membrane preparations containing EGFR can bind to GM3-coated surfaces. These data suggest that GM3 may interact directly with the EGFR. In this study, the interaction of gangliosides with the extracellular domain (ECD) of the EGFR was investigated. The purified human recombinant ECD from insect cells bound directly to ganglioside GM3. The ganglioside interaction site appears to be distinct from the EGF-binding site. In agreement with previous reports on the effects of specific gangliosides on EGFR kinase activity, the ECD preferentially interacted with GM3. The order of relative binding of other gangliosides investigated was as follows: GM3 GM2, GD3, GM4 > GM1, GD1a, GD1b, GT1b, GD2, GQ1b > lactosylceramide. These data suggest that NeuAc-lactose is essential for binding and that any sugar substitution reduces binding. In agreement with the specificity of soluble ECD binding to gangliosides, GM3 specifically inhibited EGFR autophosphorylation. Identification of a ganglioside interaction site on the ECD of the EGFR is consistent with the hypothesis that endogenous GM3 may function as a direct modulator of EGFR activity. 相似文献
16.
Gangliosides have generally been assumed to be localized primarily in the plasma membrane. Analysis of gangliosides from isolated subcellular membrane fractions of rat liver indicated that 76% of the total ganglioside sialic acid was present in the plasma membrane. Mitochondria and endoplasmic reticulum fractions, while containing only low levels of gangliosides on a protein basis, each contained approx. 10% of total ganglioside sialic acid. Gangliosides also were present in the Golgi apparatus and nuclear membrane fractions, and soluble gangliosides were in the supernatant. Individual gangliosides were non-homogeneously distributed and each membrane fraction was characterized by a unique ganglioside composition. Plasma membrane contained only 14 and 28% of the total GD1a and GD3, respectively, but 80-90% of the GM1, GD1b, GT1b and GQ1b. Endoplasmic reticulum, when corrected for plasma membrane contamination, contained only trace amounts of GM1, GD1b, GT1b and GQ1b, but 11 and 5% of the total GD1a and GD3, respectively. The ganglioside composition of highly purified endoplasmic reticulum was similar. Ganglioside biosynthetic enzymes were concentrated in the Golgi apparatus. However, low levels of these enzymes were present in the highly purified endoplasmic reticulum fractions. Pulse-chase experiments with [3H]galactose revealed that total gangliosides were labeled first in the Golgi apparatus, mitochondria and supernatant within 10 min. Labeled gangliosides were next observed at 30 min in the endoplasmic reticulum, plasma membrane and nuclear membrane fractions. Analysis of the individual gangliosides also revealed that GM3, GM1, GD1a and GD1b were labeled first in the Golgi apparatus at 10 min. These studies indicate that gangliosides synthesized in the Golgi apparatus may be transported not only to the plasma membrane, but to the endoplasmic reticulum and to other internal endomembranes as well. 相似文献
17.
Brodsky VY Zvezdina ND Nechaeva NV Avdonin PV Novikova TE Gvasava IG Fateeva VI Malchenko LA 《Cell biology international》2003,27(12):965-976
Ultradian protein synthesis rhythm was used as a marker of cell cooperation in synchronous dense and non-synchronous sparse hepatocyte cultures. Phenylephrine (2 microM, 2 min), an alpha (1)-adrenoreceptor agonist, which exerts [Ca(2+)](cyt)elevation from intracellular stores, affected protein synthesis rhythm in sparse cultures, i.e. initiated cooperative activity of the cells. The same effect was produced by 2,5-di(tertiary-butyl)-1,4-benzohydroquinone (10 microM, 2 min), which increases [Ca(2+)](cyt)by a non-receptor pathway. Pretreatment of dense cultures with the intracellular calcium chelator, 1,2-bis (2-aminophenoxy) ethane-N,N,N',N'- tetraacetic acid (acetoxymethyl) ester (BAPTA-AM) at 10-20 microM for, 30-60 min resulted in loss of the rhythm of protein synthesis, i.e. loss of cooperative activity between the cells. The medium conditioned by control dense cultures initiated rhythm in sparse cultures, whereas the conditioned medium of cultures pretreated with BAPTA-AM did not. [Ca(2+)](cyt)increase is known to occur with monosialoganglioside GM1 treatment. By ELISA estimation, the GM1 content in 3 h conditioned medium was similar in control dense cultures to that in cultures pretreated with BAPTA-AM. Bearing in mind data on the Ca(2+)-dependence of vesicle formation and shedding, the conditioned medium was separated by 150000 g centrifugation to supernatant containing monomers and micelles, and a pellet containing vesicular form of gangliosides. Only the latter initiated cooperative activity of the cells of sparse cultures. These cultures were also synchronized by GM1-containing liposomes at lower concentrations than added free GM1, 0.0003 and 0.06 microM respectively. Thus, GM1 and calcium are both involved in cell-cell synchronization. Activation of gangliosides, including GM1 and elevation of [Ca(2+)](cyt,)is known to lead to changes of protein kinase activity and protein phosphorylation resulting in modulation of oscillations in protein metabolism. 相似文献
18.
Gangliosides were recently shown to bind to calmodulin (Higashi, H., Omori, A., and Yamagata, T. (1992) J. Biol. Chem. 267, 9831-9838). This prompted us to investigate the effects of gangliosides on the calmodulin-dependent enzyme, cyclic nucleotide phosphodiesterase. Several species of gangliosides competitively inhibited calmodulin-stimulated phosphodiesterase activity, with GD1b, GT1b, and GD1a being noted to do so particularly (group 1). GM1, GQ1b, and GM2 (group 2) were less inhibitory, and GM3, GM3(NeuGc), GalCer, sulfatide, GgOse4Cer, and oligosaccharide portions of inhibitory gangliosides showed no inhibition in accordance with the binding specificity of calmodulin to gangliosides. Trypsin-activated phosphodiesterase was inhibited by gangliosides with similar specificity, indicating interactions of gangliosides with the enzyme. Inhibition, however, was less than that of calmodulin-dependent activity by these compounds and, in both cases, was eliminated by excess calmodulin. In the absence of calmodulin, group 1 gangliosides at lower concentrations activated the intact enzyme but inhibited it over a certain range of increase in concentration. Ganglioside-dependent modulation of calmodulin-dependent phosphodiesterase activity is thus shown to be due to interactions of gangliosides with both calmodulin and the enzyme, and consequently, ganglioside-calmodulin binding is likely the mechanism for regulation of the enzyme. 相似文献
19.
Abstract— The ganglioside composition of the brain of a patient with Tay-Sachs disease (TS-brain) was determined by a newly developed ganglioside-mapping procedure and compared with that of an age-matched control brain. GM2 ganglioside was the predominant component in TS-brain and the following gangliosides were also found, GM1, GD1a, GD1b and GT1 (major gangliosides in normal brain), and GM3, GD3, GD2 and GD1a-GAN (minor or undetectable components of normal brain). Individual gangliosides were isolated by column chromatography using a combination of DEAE-Sepharose, Iatrobeads and Silica Gel 60 and their structures were confirmed by comparing them with authentic standards using TLC, analysing their carbohydrate compositions by gas-liquid chromatography and cleaving them sequentially with glycosidases. The amounts of individual components were measured by quantitative densitometric scanning of the thin-layer plates. As a reflection of myelin breakdown, no sialosylgalactosyl ceramide was detectable in TS-brain. Although the total amounts of all gangliosides except GM2 in TS-brain were low, there were normal molar ratios of the main gangliosides in normal brain, that is, GM1, GD1a, GD1b and GT1. In comparison with the amount of GDla ganglioside, the amounts of GM2, GD2 and GD1a-GAN, which contain N-acetylgalactosamine as a terminal carbohydrate residue, were all elevated in TS-brain. The long chain bases of individual gangliosides contained both C-18 and C-20 sphingosine in different ratios and the ratio of C-20 to C-18 increased in the gangliosides in the order: GM2 < GM1 < GD1a < GD1a-GAN < GD1b < GT1 in both normal brain and TS-brain. In contrast, GD2 and GD3 gangliosides consisted mainly of C-18 sphingosine. The C-20 to C-18 ratios of individual gangliosides in the TS-brain were lower than those of age-matched control brain. Hexosaminidase from Turbo cornutus showed the same specific activity and Km value in catalysing the cleavage of terminal N-acetylgalactosaminyl residues from GM2, GD2 and GD1a-GAN, suggesting that the brain gangliosides that increase in Tay-Sachs disease may be cleaved by the same enzyme. 相似文献
20.
Ganglioside-modulated protein phosphorylation in myelin 总被引:5,自引:0,他引:5
K F Chan 《The Journal of biological chemistry》1987,262(5):2415-2422
Gangliosides have profound effects on the phosphorylation of several proteins in myelin. Addition of polysialogangliosides to purified guinea pig brain myelin enhanced the endogenous phosphorylation of a 62-kDa phosphoprotein, but completely inhibited the phosphorylation of myelin basic protein (MBP) (18.5 kDa). The ganglioside-stimulated phosphorylation of the 62-kDa protein was dose-dependent and -specific. Asialo-GM1, ceramide trihexosides, N-acetylneuraminic acid, or colominic acid alone could not mimic this effect, suggesting that the activation process requires both the hydrophobic head group and the anionic character of the gangliosides. Studies on the time course of this reaction revealed that it was a rapid and reversible process and was affected only very slightly by Ca2+. Thus, the stimulatory effect of gangliosides may not involve Ca2+-gangliosides complexes or proteolysis, but may be mediated through an activation of a ganglioside-dependent protein kinase or due to substrate protein-glycolipid interaction. Modulation of the phosphorylation of MBP by gangliosides varies with the states of phosphorylation of this protein. Prior addition of ganglioside to myelin inhibited the phosphorylation of MBP. However, addition of gangliosides to myelin subsequent to maximal phosphorylation of MBP retarded the dephosphorylation of this protein. Phosphorylation of isolated MBP by protein kinase C was stimulated by gangliosides, provided phosphatidylserine was present. In contrast, the glycolipid inhibited the phosphorylation of a unique site catalyzed by cAMP-dependent protein kinase. This site was distinct from those phosphorylated by protein kinase C and was also sensitive to chymotryptic cleavage. Although the exact physiological significance of protein phosphorylation in myelin has yet to be established, gangliosides may play an important role in the modulation of this reversible post-translational modification mechanism. 相似文献