Journal of cyclic nucleotide and protein phosphorylation research最新文献_第7页

Adipocyte cyclic nucleotide phosphodiesterase activation by vanadate. 钒酸盐活化脂肪细胞环核苷酸磷酸二酯酶。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

J E Souness, W J Thompson, S J Strada

{"title":"Adipocyte cyclic nucleotide phosphodiesterase activation by vanadate.","authors":"J E Souness, W J Thompson, S J Strada","doi":"","DOIUrl":"","url":null,"abstract":"The addition of vanadate (Na3VO4) to intact isolated rat adipocytes stimulated cAMP phosphodiesterase activity (Type IV) in the particulate (P2) fraction. Vanadate increased the Vmax of the Type IV phosphodiesterase activity without affecting its apparent substrate affinity. Na3VO4 also stimulated cAMP hydrolysis of cell-free particulate and cytosolic fractions, but this activation required the presence of reduced glutathione (GSH). The mixture of vanadate and glutathione appeared as an emerald green solution (V-GSH complex), which was shown by EPR to contain vanadyl ion. No effect of either GSH or Na3VO4 alone on cell-free particulate cAMP phosphodiesterase activity was observed; however, Na3VO4, alone or in combination with GSH, stimulated cGMP hydrolysis in this subcellular fraction. The V-GSH complex increased the Vmax of the particulate cAMP phosphodiesterase activity without affecting its apparent Km. The activating effect of the complex was rapid in onset, persistent over 30 minutes, and reversible. The EC50 for activation of the particulate cAMP phosphodiesterase was approximately 5 microM Na3VO4 (maintaining the GSH:Na3VO4 molar ratio at 2:1); maximal stimulation was achieved at 0.1 mM Na3VO4. Purified microsomal membranes showed activation similar to that of the P2 fraction, while only a 60% stimulation was observed in purified plasma membranes. The V-GSH complex increased basal insulin-activated Type IV phosphodiesterase activity to a common maximal level. Detergent-solubilized cAMP-phosphodiesterase from the P2 fraction was stimulated 2.5-fold by the V-GSH complex. Limited trypsin treatment of P2 membranes activated cAMP phosphodiesterase and abolished the stimulatory effect of the V-GSH complex. These results are generally consistent with the hypothesis that V-GSH complex activates Type IV phosphodiesterase by an indirect mechanism, which appears to involve predominantly membrane bound components that may be biologically important enzyme regulatory elements.","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"14126846","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Forskolin modulates cyclic AMP generation in the rat myometrium. Interactions with isoproterenol and prostaglandins E2 and I2. 福斯克林调节大鼠肌层循环AMP的生成。与异丙肾上腺素和前列腺素E2和I2的相互作用。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

A Mokhtari, L Do Khac, Z Tanfin, S Harbon

{"title":"Forskolin modulates cyclic AMP generation in the rat myometrium. Interactions with isoproterenol and prostaglandins E2 and I2.","authors":"A Mokhtari, L Do Khac, Z Tanfin, S Harbon","doi":"","DOIUrl":"","url":null,"abstract":"In the intact rat myometrium, forskolin stimulated cyclic AMP generation and markedly potentiated increases in cyclic AMP caused by isoproterenol, prostaglandin E2 and prostacyclin. The diterpene increased the maximal responses and lowered the EC50 for both isoproterenol- and prostaglandin-stimulated cyclic AMP accumulation. Forskolin did not modify the Ki for the beta-adrenergic antagonist propranolol. Activation of cyclic AMP generation by forskolin was biphasic with respect to concentration; the major response being mediated by a low affinity interaction (Kapp 28 microM) and a minor effect being due to an interaction with a high affinity site (Kapp 0.5 microM). By contrast, the synergistic effect of the diterpene with isoproterenol, prostaglandin E2 as well as with cholera toxin, involved a single component of high affinity (Kapp 0.5 to 2 microM), which was thus considered to be associated with the activated complex of the cyclase catalytic subunit and the guanine nucleotide regulatory protein. Forskolin could further partially maintain isoproterenol-mediated synergism in a beta-adrenergic desensitized tissue. In myometrial membrane preparations, forskolin stimulated adenylate cyclase activity but failed to potentiate isoproterenol- and prostaglandin E2-mediated activation.","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"14125001","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Determination of cyclic AMP-dependent protein kinase subunits by an immunoassay reveals a different subcellular distribution of the enzyme in rat parotid than does determination of the enzyme activity. 通过免疫测定法测定环amp依赖性蛋白激酶亚基，揭示了该酶在大鼠腮腺中的亚细胞分布与测定酶活性不同。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

G Schwoch, S M Lohmann, U Walter, U Jung

{"title":"Determination of cyclic AMP-dependent protein kinase subunits by an immunoassay reveals a different subcellular distribution of the enzyme in rat parotid than does determination of the enzyme activity.","authors":"G Schwoch, S M Lohmann, U Walter, U Jung","doi":"","DOIUrl":"","url":null,"abstract":"The distribution of cyclic cAMP-dependent protein kinase in subcellular fractions of rat parotid was determined by two independent biochemical methods, measurement of kinase catalytic activity or by quantitation of the catalytic and regulatory subunits in an enzyme-linked immunosorbent assay using monospecific antibodies. The major amount (85%) of the catalytic activity was found associated with the 100,000 g soluble fraction, whereas only 1/3 of the total catalytic subunit was demonstrated in the soluble fraction by the immunoassay. The immunoassay results furthermore indicated that approximately 50% of the total cellular protein kinase was associated with the extranuclear particulate fraction and that the predominant form of the kinase in the particulate fractions was the type II isoenzyme. The reasons for the differences in the distribution of the protein kinase demonstrated by the two methods were examined. Incomplete extraction of membrane-bound protein kinase and the influence of membrane localized ATPases on activity measurements were, at least in part, responsible for the low percentage of kinase activity measured in the particulate fractions. These results emphasize that the precise quantitation of protein kinase subunits merits investigation by more than one method. For the parotid, the finding that approximately 2/3 of the total catalytic subunit may be particulate associated provides additional evidence that cyclic AMP could be involved in membrane mechanisms of hormone-regulated secretion.","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"14125003","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Adenylate cyclases in two populations of membranes purified from neuroblastoma X glioma hybrid (NG 108-15) cells. 从神经母细胞瘤X胶质瘤杂交(NG 108-15)细胞纯化的两种膜中的腺苷酸环化酶。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

F W Sweat, W A Klee

{"title":"Adenylate cyclases in two populations of membranes purified from neuroblastoma X glioma hybrid (NG 108-15) cells.","authors":"F W Sweat, W A Klee","doi":"","DOIUrl":"","url":null,"abstract":"Membranes from neuroblastoma X glioma NG108-15 hybrid cells were purified by equilibrium centrifugation on continuous and discontinuous gradients of sorbitol, using homogenates of cells which were pretreated with concanavalin A to increase membrane density. Adenylate cyclase was purified 24-fold in a heavy (H) membrane fraction from discontinuous gradients, opiate-stimulated guanosine-5'-triphosphatase was purified 3-fold, and opiate binding to receptors was increased 10-fold in this fraction. The relative purification of this membrane fraction is also verified by the fact that it contains a single protein (Mr = 58,000) which is covalently labeled by a reactive opiate analog (Klee, W. A., Simonds, W. F., Sweat, F. W., Burke, T. R., Jacobson, A. E., and Rice, K. C. (1982) FEBS Lett. 150, 125). The method of plasma membrane purification after cell treatment with concanavalin A (Lutton, J. K., Frederich, R. C. Jr., and Perkins, J. P. (1979) J. Biol. Chem. 254, 11181) appears generally applicable as established here with 3H-concanavalin A. Between 15 and 20% of the adenylate cyclase in whole-cell homogenates was recovered at low densities in continuous and discontinuous gradients and was only purified 2-fold above activity in the cell homogenate. There are significant differences between ligand binding, adenylate cyclase, and GTPase activities in light (L) and heavy (H) membrane fractions. GTPase activity in the L-membrane fraction was decreased from that in the cell homogenate and was not stimulated by opiates. Adenylate cyclase from L-membranes is only slightly inhibited by opiates in support of other data relating opiate inhibition to stimulation of GTPase (Koski, G., and Klee, W. A. (1981) Proc. Natl. Acad. Sci. 78, 4185).","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"15198659","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

The resolution of agonist alpha 2-adrenergic receptor complexes from unoccupied receptors or antagonist-alpha 2 receptor complexes using DEAE chromatography. 用DEAE色谱法从未占用受体或拮抗剂α 2受体复合物中分离激动剂α 2-肾上腺素能受体复合物。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

L E Limbird, S T MacMillan, D L Kalinoski

引用次数: 0

Intracellular injection of cyclic GMP depresses cardiac slow action potentials. 细胞内注射环GMP可抑制心脏慢动作电位。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

G M Wahler, N Sperelakis

{"title":"Intracellular injection of cyclic GMP depresses cardiac slow action potentials.","authors":"G M Wahler, N Sperelakis","doi":"","DOIUrl":"","url":null,"abstract":"Our laboratory has shown that intracellular injection of cyclic AMP (cAMP) transiently enhances slow APs in myocardial cells, presumably by phosphorylating slow channels. To test if cGMP also plays a role in cardiac slow channel function, superfusion with 8-Br-cGMP, and intracellular injections of cGMP were carried out in guinea pig papillary muscles (stimulated at 0.5 Hz at 37 degrees C). In normal (4.7 mM K+) Tyrode's solution, 0.1 mM 8-Br-cGMP depressed contractions and had variable effects on the duration of the fast APs. Slow APs were elicited by electrical stimulation (in 25 mM K+-Tyrode's solution) following the addition of 10 mM TEA and doubling the bath [Ca] (to 4.0 mM) or addition of 0.2 microM isoproterenol. Slow APs are dependent on the slow inward current carried through voltage- and time-dependent slow channels. 8-Br-cGMP (0.1 microM - 1 mM) superfusion depressed or abolished slow APs and accompanying contractions. cGMP (5-100 mM Na+ salt in 0.2 M KC1) was injected by application of pressure pulses (40-75 psi, 1-30 sec duration) to the recording microelectrode. cGMP injection transiently depressed (n = 15) or abolished (n = 4) the slow APs. The effect began 1 min after the onset of the pulse, reached a maximum at 2 min and recovered fully within 5-6 min. Thus, it appears that the intracellular cGMP level can modulate the slow inward current in a direction opposite to that of cAMP. These effects may both be due to cyclic nucleotide-dependent phosphorylations.","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"14117714","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Effects of thyroidectomy, insulin, and phospholipids on cyclic AMP phosphodiesterase in rat adipocyte plasma membranes. 甲状腺切除术、胰岛素和磷脂对大鼠脂肪细胞膜环AMP磷酸二酯酶的影响。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

C Correze, H Thibout

{"title":"Effects of thyroidectomy, insulin, and phospholipids on cyclic AMP phosphodiesterase in rat adipocyte plasma membranes.","authors":"C Correze, H Thibout","doi":"","DOIUrl":"","url":null,"abstract":"The effect of thyroid status on the cyclic nucleotide phosphodiesterase in adipocyte plasma membranes has been studied. In euthyroid rat fat cells, about 7% of the total cAMP phosphodiesterase was found in the plasma membrane. Thyroidectomy doubled both the enzyme's total activity in this fraction, and its specific activity (60 versus 37 pmol/min/mg) compared to euthyroid rat plasma membrane. In membranes from thyroidectomized rats, phosphodiesterase hydrolyzed cAMP with a single Km of 2 microM, whereas in euthyroid rat membranes, Lineweaver Burk plots were non-linear, with apparent Kms of 0.5 and 5 microM. This phosphodiesterase activity was insensitive to exogenous guanine nucleotides and calcium. In vivo injection of triiodothyronine restored phosphodiesterase activity in plasma membranes from thyroidectomized rats to the values obtained for euthyroid rats. Centrifugation on a 10 to 45% sucrose density gradient of the plasma membrane fractions gave two main peaks of phosphodiesterase activity which hydrolyzed the cAMP in adipocyte plasma membranes from both euthyroid and thyroidectomized rats. The distribution profiles for these activities were very similar in the two plasma membrane preparations. The peaks of phosphodiesterase and 5'nucleotidase activity coincided. Thyroidectomy raised the phosphodiesterase activity of these two peaks, particularly of the first. The cAMP phosphodiesterase activities in both hypothyroid and control plasma membrane preparations were also sensitive to insulin and were activated by phospholipase A2 and three anionic phospholipids. Thyroid hormones therefore regulate the degradation of cAMP in plasma membranes by a mechanism which seems different from the one involved in the action of insulin, and is independent of the membrane phospholipid composition.","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"14120828","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A monoclonal antibody to a membrane component that interacts with the beta-adrenergic receptor. 与-肾上腺素能受体相互作用的膜成分的单克隆抗体。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

D M Chuang

{"title":"A monoclonal antibody to a membrane component that interacts with the beta-adrenergic receptor.","authors":"D M Chuang","doi":"","DOIUrl":"","url":null,"abstract":"A monoclonal antibody has been obtained using a combination of in vivo and in vitro immunization with a digitonin extract of purified plasma membranes from frog erythrocytes. This antibody was found to immunoprecipitate a fraction of solubilized beta-adrenergic receptor (labeled with 125I-iodohydroxybenzylpindolol) derived from frog erythrocytes and a few other sources. This immunoglobulin also significantly activated adenylate cyclase in isolated erythrocyte plasma membranes measured in the presence of GTP or GTP plus isoproterenol. Immunoprecipitation of labeled erythrocyte surface proteins and immunoblotting of the digitonin-extract of erythrocyte plasma membrane revealed that the antibody interacted with a protein with a Mr = 43,000 and pI = 6.2 32P-ADP-ribosylated alpha subunit of Ns (with Mr = 44,000) solubilized from frog erythrocyte membranes failed to be immunoprecipitated by the antibody. Thus the antigenic protein is distinct from the beta-adrenergic binding site and alpha subunit of Ns and therefore may be an unidentified component of the beta-adrenergic receptor-adenylate cyclase system. This monoclonal antibody may be a useful tool for future studies for topological and functional interactions between the beta-adrenergic receptor and other membrane components.","PeriodicalId":15406,"journal":{"name":"Journal of cyclic nucleotide and protein phosphorylation research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"1985-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"14126179","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

The binding of cyclic nucleotide analogs to a purified cyclic GMP-stimulated phosphodiesterase from bovine adrenal tissue. 环核苷酸类似物与纯化的环gmp刺激的牛肾上腺组织磷酸二酯酶的结合。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

C Erneux, F Miot, P J Van Haastert, B Jastorff

引用次数: 0

Tribute to Shiro Kakiuchi 1929-1984. 1929-1984年向角内四郎致敬。

Journal of cyclic nucleotide and protein phosphorylation research Pub Date : 1985-01-01

H Hidaka

引用次数: 0