Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Collagenase preparations (a mixture of enzymes including collagenase, clostripain, and a casein-degrading protease) degraded the beta subunit (Mr = 95,000) of the purified insulin receptor into fragments of Mr less than 15,000, without degrading the alpha subunit. The resulting beta-digested insulin receptor preparations were found to bind insulin as well as control insulin receptor, as assessed by either cross-linking of 125I-insulin to the digested receptor or by separating insulin bound to receptor from free insulin by high performance liquid chromatography. Moreover, the beta-digested insulin receptor preparations were still precipitated by a monoclonal antibody directed against the insulin-binding site. In contrast, the beta-digested insulin receptor lacked protein kinase activity since it no longer phosphorylated either itself, or an exogenous substrate, calf thymus histone. These results support the identification of the beta subunit of the insulin receptor as a protein kinase.
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PMID:Preferential degradation of the beta subunit of purified insulin receptor. Effect on insulin binding and protein kinase activities of the receptor. 631 28

Protein I has been identified and compared in membranes prepared from chick and rat forebrain. Based upon five criteria known to characterize protein I, namely, (1) its ability to serve as a substrate for both the cyclic AMP-dependent protein kinase and (2) the Ca2+- dependent, calmodulin-requiring protein kinase, (3) its ability to be extracted from membranes at low pH, (4) its characteristic pattern of digestion by collagenase, and (5) its existence as a basic protein, we have determined that although protein I of rat brain consists of the usual doublet polypeptides Ia and Ib, only a single chick forebrain polypeptide is detectable which possesses protein I-like properties.
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PMID:Identification and comparison of protein I in chick and rat forebrain. 631 4

Synapsin I (protein I) is a neuron-specific phosphoprotein, which is a substrate for cAMP-dependent and Ca/calmodulin-dependent protein kinases. In two accompanying studies (De Camilli, P., R. Cameron, and P. Greengard, and De Camilli, P., S. M. Harris, Jr., W. B. Huttner, and P. Greengard, 1983, J. Cell Biol. 96:1337-1354 and 1355-1373) we have shown, by immunocytochemical techniques at the light microscopic and electron microscopic levels, that synapsin I is present in the majority of, and possibly in all, nerve terminals, where it is primarily associated with synaptic vesicles. In the present study we have prepared a highly purified synaptic vesicle fraction from rat brain by a procedure that involves permeation chromatography on controlled-pore glass as a final purification step. Using immunological methods, synapsin I concentrations were determined in various subcellular fractions obtained in the course of vesicle purification. Synapsin I was found to copurify with synaptic vesicles and to represent approximately 6% of the total protein in the highly purified synaptic vesicle fraction. The copurification of synapsin I with synaptic vesicles was dependent on the use of low ionic strength media throughout the purification. Synapsin I was released into the soluble phase by increased ionic strength at neutral pH, but not by nonionic detergents. The highly purified synaptic vesicle fraction contained a calcium-dependent protein kinase that phosphorylated endogenous synapsin I in its collagenase-sensitive tail region. The phosphorylation of this region appeared to facilitate the dissociation of synapsin I from synaptic vesicles under the experimental conditions used.
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PMID:Synapsin I (protein I), a nerve terminal-specific phosphoprotein. III. Its association with synaptic vesicles studied in a highly purified synaptic vesicle preparation. 640 12

Certain metabolic properties of hormonally responsive osteogenic sarcoma cells derived from a transplantable rat tumor have been compared with those of related normal rat bone cells. All studies were carried out on cells grown in monolayer culture. Normal rat bone cells derived by repeated collagenase/trypsin digestion of newborn rat calvaria. Bone cells selected for comparison were thought to be osteoblast-like, as judged by enrichment of alkaline phosphatase and adenylate cyclase responsiveness to parathyroid hormone and prostaglandin E2. The adenylate cyclases of the two cell strains were similarly stimulated by a range of prostanoids and their metabolites and analogs. Morphology showed the two cell strains to be similar; the only obvious difference was a multilayering of cells in the sarcoma cultures, while the normal cultures showed abundant extracellular fibril formation which was not seen in the tumor cells. Investigation of the cAMP-dependent protein kinase isoenzymes showed the presence of two forms in both cell types, one eluting at a low salt concentration and the other at a high salt concentration. There was approximately twice the amount of the first isoenzyme compared to the second isoenzyme. The results indicate the usefulness of the two cell strains to elucidate further the molecular mechanisms of action of parathyroid hormone and prostaglandins.
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PMID:Functional properties of hormonally responsive cultured normal and malignant rat osteoblastic cells. 693 60

TNF stimulated transcription and secretion of the metalloproteinases collagenase and stromelysin in porcine articular chondrocytes. TNF induced metalloproteinase transcription could be inhibited with either protein kinase inhibitors (H7 or staurosporine) or by raising intracellular cAMP levels. HA1004, a protein kinase inhibitor structurally related to H7 but with a higher Ki for protein kinase C had no effect on TNF induced message levels. TNF treatment of chondrocytes did not induce membrane associated PKC or increase intracellular cAMP levels. Our results are consistent with the involvement of a staurosporine and H7 sensitive protein kinase distinct from PKC in TNF signal transduction in chondrocytes.
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PMID:Protein kinase regulation of tumor necrosis factor alpha stimulated collagenase and stromelysin message levels in chondrocytes. 750 65

We evaluated the role of protein phosphorylation in cAMP-mediated amylase exocytosis from parotid acinar cells by using H89, a new protein kinase A (PKA) inhibitor, which is more lipophilic and 25 times more potent than H8. In our previous studies, H8 markedly inhibited protein phosphorylation without decreasing amylase release [Takuma, T. (1988) Biochem. J. 256, 867-871]. These findings were completely reproduced even in the small acini that were prepared by trypsin treatment before collagenase digestion. In the present study, however, H89 strongly inhibited both amylase release and protein phosphorylation in a dose-dependent manner. The inhibitory effect was specific for PKA at least up to 33 microM, since 33 microM H89 did not block amylase release stimulated by PMA. H85, a closely related compound of H89 without inhibitory effect on PKA, did not prevent amylase release or protein phosphorylation at least up to 33 microM. These results suggest that protein phosphorylation by PKA is involved in cAMP-mediated amylase exocytosis. The inhibition of protein phosphorylation by H8 might be insufficient or inadequate for blocking of amylase release.
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PMID:Evidence for the involvement of protein phosphorylation in cyclic AMP-mediated amylase exocytosis from parotid acinar cells. 750 58

Galanin has numerous effects on gastrointestinal motility in different species; however, its cellular basis of action in mediating these effects is unclear. Dispersed gastric smooth muscle cells have been shown to possess high-affinity galanin receptors that increase cAMP and cause relaxation. Recent studies show some smooth muscle relaxants such as VIP cause relaxation by both cAMP-dependent and -independent mechanisms. It is unknown if galanin's cellular basis of relaxation is similar or different from that of VIP. To investigate galanin's relaxant effect and compare it to VIP's effect, dispersed smooth muscle cells from guinea pig stomach were prepared by collagenase digestion. The mean length in resting cells was 110 +/- 2 microns and, with carbachol treatment, contracted to 89 +/- 2 microns. VIP and galanin alone had no effect on cell length, but each caused a dose-dependent inhibition of carbachol-induced contraction and both had an EC50 of 3-7 nM. Galanin (1 microM) and VIP (1 microM) increased cellular cAMP from 118 +/- 10 pmol/10(6) cells in control to 212 +/- 14 and 214 +/- 12 pmol/10(6) cells, respectively. The protein kinase A inhibitor, Rp-cAMPS, at 100 microM, completely inhibited the relaxant effect of an EC50 concentration of galanin (3 nM), but only inhibited that by VIP by 80% (p < 0.05). Adding the nitric oxide inhibitor, L-NNA (NG-nitro-L-arginine), at 100 microM did not alter the length of resting cells or inhibit carbachol-induced contraction. However, L-NNA (100 microM) decreased VIP-induced relaxation by 45%, whereas it had no effect on galanin-induced relaxation.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Galanin-induced relaxation in gastric smooth muscle cells is mediated by cyclic AMP. 753 25

We examined the common signal transduction mechanisms governing collagenase (MMP-1), stromelysin-1 (MMP-3), and tissue inhibitor of metalloproteases (TIMP-1) gene expression in human synovial fibroblasts for insight into the pathophysiology of arthritis. MMP-1, MMP-3, and TIMP-1 expression and synthesis were induced in cultured human synoviocytes with recombinant human interleukin 1 beta in the absence or presence of either chemical inhibitors of protein kinase A and C (PKA, PKC), or prostaglandin E2, or cyclic AMP (cAMP) mimetics. We used enzyme immunoassays (EIA) to determine MMP-1, MMP-3, and TIMP-1 antigen levels in spent culture medium and Northern hybridization to measure steady state mRNA expression levels. Extracellular signals (e.g., IL-1, phorbol myristic acetate) that result in the activation of cytoplasmic PKC augment in tandem the expression and synthesis of MMP-1, MMP-3, and TIMP-1 in human synovial fibroblasts. In addition, such signals induce nuclear transcription factors (e.g., activator protein 1) that bind to common gene regulatory elements and augment promoter activity of MMP-1, MMP-3, and TIMP-1 gene promoter constructs. In contrast, signals that activate PKA oppose PKC mediated signals, in that the expression of MMP-1, MMP-3, and TIMP-1 are suppressed. Experimental data suggest that the expression of MMP-1, MMP-3, and TIMP-1 are coordinated through a series of common cytoplasmic signal transducing pathways, cis regulatory elements, and nuclear trans acting factors.
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PMID:Coordinate regulation of matrix metalloproteases and tissue inhibitor of metalloproteinase expression in human synovial fibroblasts. 775 15

Osteocyte-like cells were prepared by sequentially treating calvaria from newborn rats with collagenase and chelating agents. On a reconstituted gel of basement membrane components, cells from the third collagenase digest displayed a round shape and expressed the highest level of alkaline phosphatase with minimal osteocalcin deposition into the matrix. On the other hand, cells derived from the interior after EDTA treatment exhibited well-developed dendritic cell processes and expressed essentially no alkaline phosphatase. The latter population also showed quite distinct characteristics such as higher extracellular activities of casein kinase II and ecto-5'-nucleotidase and the extracellular accumulation of a large amount of osteocalcin associated with mineral. These diverse phenotypic and protein expressions as well as the sites from which each population of cells were recovered strongly suggest that we have isolated osteoblastic and osteocytic cells. Bone sialoprotein II was extracellularly phosphorylated by casein kinase II in osteocytic cells but not in osteoblastic cells. We discuss the possibility that differentiation of young osteocytes from osteoblasts may facilitate the biochemical sequence of mineral deposition in the bone matrix.
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PMID:Matrix mineralization and the differentiation of osteocyte-like cells in culture. 775 2

Prostaglandins (PGs) may stimulate or inhibit bone cell replication and protein synthesis. These disparities may be concentration or time dependent, or occur in discrete cell types or by different second signals. Cell populations that express progressive degrees of osteoblast-like activity can be obtained by serial collagenase digestion of fetal rat parietal bone. The first (population 1) appears less differentiated, whereas the later (populations 3-5) exhibit biochemical features characteristic of osteoblasts. Within 24 h of treatment, three separate PGs increased DNA synthesis in population 1 with relative potencies of PGE1 < PGE2 < PGF2 alpha. By contrast, PGE1 and PGE2 (both strong cAMP inducers) inhibited basal DNA synthesis in population 3-5. These differences were paralleled by analogous changes in collagen and noncollagen synthesis in each population. The mitogenic effect in population 1 persisted for 72 h, and at later times was sensitive to indomethacin. These changes were unlikely to be cAMP dependent, as PGF2 alpha did not induce cAMP production, and the cAMP inducer forskolin was inhibitory. Moreover, phorbol ester treatment enhanced DNA synthesis to a greater extent in population 1 than in populations 3-5, and cotreatment with H-8 (at Km, approximately 10 microM) and staurosporine (at Km, approximately 0.01 microM) decreased the mitogenic effect of PGs in population 1, consistent with a reduction in protein kinase-C activation. These studies suggest that PGs activate less differentiated bone cells by a protein kinase-dependent event, whereas cAMP (induced by PGE1 and PGE2) decreases DNA and protein synthesis in more differentiated bone cells and tempers the increase in cellular activation found in population 1. Consequently, agents or events that increase the synthesis of specific PGs could differentially regulate, in positive and negative ways, biochemical activities in discrete bone cell populations.
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PMID:Differential actions of prostaglandins in separate cell populations from fetal rat bone. 792 24


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