Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Calpactins I and II are proteins that bind Ca2+, phospholipids, actin and spectrin; they are also major substrates of oncogene and growth-factor-receptor tyrosine kinases. Since calpactins have been proposed to provide a link between membrane lipids and the cytoskeleton, we examined in detail the interactions between purified calpactin I and phospholipid liposomes. We focused on the Ca2+-dependence, the effects of phosphorylation of calpactin I by p60v-src (the protein kinase coded for by the Rous-sarcoma-virus oncogene), and the effects of the binding of calpactin I light chain to calpactin I heavy chain. Binding of the light chain to the heavy chain increased the affinity of calpactin I for phosphatidylserine (PS) liposomes. The opposite effect was observed for phosphorylation by p60v-src; phosphorylation decreased the affinity of calpactin I for PS liposomes. These two opposite effects appeared to be independent, since phosphorylation did not prevent light-chain binding to the heavy chain. Calpactin I was found, by the use of three different techniques, to bind to phospholipid liposomes at less than 10(-8) M free Ca2+. This result is in contrast with those of previous studies, which indicated that greater than 10(-6) M free Ca2+ was required. Our findings suggest that calpactin I may be bound to phospholipids in vivo at Ca2+ concentrations of about 1.5 x 10(-7) M, typical of resting unstimulated cells, and that this interaction may be modulated by light-chain binding and phosphorylation by p60v-src.
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PMID:Regulation of calpactin I phospholipid binding by calpactin I light-chain binding and phosphorylation by p60v-src. 296 67

The mos oncogene of Moloney murine sarcoma virus encodes a protein of approximately 37,000 daltons (designated p37mos). We have detected a serine protein kinase activity which is closely associated with p37mos in immune complexes obtained with antibodies [anti-mos(37-55) serum] that were generated with a peptide containing amino acids 37 through 55 of the v-mos protein (S. A. Maxwell and R. B. Arlinghaus, Virology 143:321-333, 1985). Immune complexes that were derived with antibodies generated against peptides representing the C-terminal 8 or 12 amino acids of v-mos (anti-C2 and anti-C3 serum, respectively) exhibited very little kinase activity capable of phosphorylating p37mos. Treatment of anti-mos(37-55) complexes containing active v-mos kinase with anti-C3 or anti-C2 serum resulted in a dramatic reduction of the in vitro phosphorylation of p37mos. Antiserum blocked with the appropriate C-terminal peptide had no inhibitory effect on the phosphorylation of p37mos in anti-mos(37-55) complexes which indicated that the inhibition of v-mos kinase activity was a specific effect of these antibodies. The specific inhibition of the in vitro phosphorylation of p37mos by antibodies directed against the C terminus of the v-mos protein provides strong evidence that the v-mos gene encodes a serine protein kinase. In addition, the extreme C terminus of p37mos may be critical for an active v-mos kinase.
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PMID:Use of site-specific antipeptide antibodies to perturb the serine kinase catalytic activity of p37mos. 299 8

Two proteins, termed P85gag-mos and P58gag, are encoded by the temperature-sensitive transformation mutant, ts110 Moloney murine sarcoma virus (MuSV). Based on temperature-shift studies, P85gag-mos is believed to be important for the transforming potential of ts110 MuSV and has been found to be associated with a thermolabile kinase activity that phosphorylates both P85gag-mos and P58gag in immune complexes. Modifications of the original kinase assay conditions are reported here that have allowed a 30-fold increase in the specific activity of P85gag-mos phosphorylated in vitro. The in vitro P85gag-mos-phosphorylating activity was found to be unresponsive to 10 microM-cAMP or 10 microM-cGMP. Addition of 1 mM-pyrophosphate, a known phosphatase inhibitor, to the reaction mixture resulted in an increased yield of phosphorylated P85gag-mos and P58gag; the molar phosphate incorporation per mole of P85gag-mos increased from 0.032 to 0.9, whereas the specific activity of in vitro-phosphorylated P58gag increased 18-fold, from 0.013 to 0.234. pH curves of the in vitro kinase reaction further confirmed the presence of phosphatase activity; in the absence of pyrophosphate, a sharp optimum at pH 4 to 5 was observed, whereas it shifted broadly to pH 7.0 in the presence of pyrophosphate. Under the latter conditions, several experiments were performed in order to determine if the kinase was associated with either gag or mos sequences of P85gag-mos. Antisera directed against p15, p12 and p30 sequences of the gag protein region of P85gag-mos yielded immune complexes that allowed phosphorylation in vitro of P85gag-mos. No phosphorylating activity was detected in immune complexes containing MuSV-124-encoded P62gag. An anti-mos serum generated against a synthetic peptide representing the predicted v-mos amino acid residues 37 to 55 recognizes P85gag-mos and allowed phosphorylation of P85gag-mos in vitro in the absence of P58gag. Peptide mapping of both phosphorylated P85gag-mos and P58gag, by using a combination of Cleveland and Western/immunoperoxidase techniques, demonstrated that P85gag-mos became phosphorylated not only on gag sequences, but also at the N-terminal portion of v-mos. Phosphoamino acid analyses of P85gag-mos and P58gag phosphorylated in vitro under these modified conditions yielded predominantly phosphoserine and lesser amounts of phosphothreonine. Metabolically 32P-labelled P85gag-mos and P58gag were also found to contain phosphoserine and phosphothreonine. Based on these results, we conclude that a cAMP-independent, serine/threonine protein kinase activity is associated with the mos sequences of P85gag-mos.
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PMID:A cAMP-independent serine/threonine kinase activity is associated with the mos sequences of ts110 Moloney murine sarcoma virus-encoded P85gag-mos. 299 51

An antiserum directed against amino acid residues 37-55 [anti-mos (37-55) serum] of the predicted v-mos sequence was used to precipitate p37mos from Moloney murine sarcoma virus-124 (Mo-MuSV-124) acutely infected 3T3 cells. Proteins with sizes ranging from p37mos to 43 kDa (p43) were found to be phosphorylated when anti-mos (37-55) immune complexes containing p37mos were incubated with [gamma-32P]ATP and Mn2+. The phosphorylation of p37mos and p43 could be specifically blocked when the anti-mos (37-55) serum was incubated with 37-55 cyclic mos peptide prior to immunoprecipitation, but not if the serum was preincubated with an unrelated peptide representing amino acids of the myc protein sequence. Anti-mos (37-55) immune complexes from uninfected 3T3 cells did not produce any phosphorylated proteins the size of p37mos or p43. However, a 50-kDa protein (p50) was phosphorylated in both unblocked and mos peptide-blocked anti-mos (37-55) immune complexes from infected 3T3 cells, and in immune complexes from uninfected cells. Quercetin, an inhibitor of some protein kinases, inhibited the kinase phosphorylating p50 but not the kinase phosphorylating p37mos and p43. Preabsorption of the cell extract prior to immunoprecipitation with an excess of formalin-fixed Staphylococcus aureus, complexed with preimmune normal rabbit serum IgG, specifically removed the kinase phosphorylating p50. The amount of in vitro phosphorylated p37mos and p43 in the immune-complex kinase assay reached a maximum in extracts of 3T3 cells 2-3 days postinfection with Mo-MuSV 124 but decreased to trace levels after 5 days. Metabolically and in vitro phosphorylated p37mos generated an identical pattern of phosphopeptides upon partial V8 protease digestion. Based on peptide mapping and a kinetic analysis of the in vitro phosphorylation reaction, p37mos appears to be a precursor to the p43 phosphorylated species. Phosphoamino acid analyses revealed only phosphoserine in in vitro phosphorylated p37mos and p43mos. It was concluded that p37mos is closely associated with a serine kinase activity and that the in vitro phosphorylation of p37mos may lead to formation of a highly modified mos protein (p43) by way of superphosphorylation.
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PMID:Serine kinase activity associated with Maloney murine sarcoma virus-124-encoded p37mos. 299 8

The transforming gene product encoded by Moloney murine sarcoma virus clone 124, p37mos, contains a lysine residue (lysine-121) that is conserved among all members of the protein kinase family. This lysine has been shown to be part of a conserved ATP-binding site in both the catalytic subunit of the cAMP-dependent protein kinase and p60v-src. We wished to determine whether this lysine is required for the transforming activity of p37mos. Two site-specific mutations were therefore constructed, which result in the substitution of an aspartic acid or arginine codon in place of the codon for lysine-121. Both mutations abolished the ability of the mos gene to transform cells. These results show that lysine-121 is required for the ability of p37mos to transform cells and provide evidence for an ATP-binding site in p37mos. Furthermore, these results suggest that the conserved lysine residue is specifically involved in the catalytic activity of protein kinases in general.
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PMID:Lysine residue 121 in the proposed ATP-binding site of the v-mos protein is required for transformation. 299 82

The P130gag-fps transforming protein of Fujinami sarcoma virus (FSV) possesses tyrosine-specific protein kinase activity and autophosphorylates at Tyr-1073. Within the kinase domain of P130gag-fps is a putative ATP-binding site containing a lysine (Lys-950) homologous to lysine residues in cAMP-dependent protein kinase and p60v-src which bind the ATP analogue p-fluorosulfonylbenzoyl-5' adenosine. FSV mutants in which the codon for Lys-950 has been changed to codons for arginine or glycine encode metabolically stable but enzymatically defective proteins which are unable to effect neoplastic transformation. Kinase-defective P130gag-fps containing arginine at residue 950 was normally phosphorylated at serine residues in vivo suggesting that this amino acid substitution has a minimal effect on protein folding and processing. The inability of arginine to substitute for lysine at residue 950 suggests that the side chain of Lys-950 is essential for P130gag-fps catalytic activity, probably by virtue of a specific interaction with ATP at the phosphotransfer active site. Tyr-1073 of the Arg-950 P130gag-fps mutant protein was not significantly autophosphorylated either in vitro or in vivo, but could be phosphorylated in trans by enzymatically active P140gag-fps. These data indicate that Tyr-1073 can be modified by intermolecular autophosphorylation.
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PMID:A lysine in the ATP-binding site of P130gag-fps is essential for protein-tyrosine kinase activity. 300 19

We describe the expression of the Moloney murine sarcoma virus env-mos protein (p37mos) in yeast under the control of the yeast GAL1 promoter. Consistent with our previous results concerning the p37mos protein kinase made in virus infected mouse cells, p37mos produced in yeast possesses autophosphorylation activity in an immune complex kinase assay using anti-mos (37-35) serum.
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PMID:Moloney murine sarcoma virus encoded p37mos expressed in yeast has protein kinase activity. 301 29

The role of prostaglandins in cellular differentiation and transformation has been widely studied. We have found previously that prostaglandin E2 production was greatly diminished in dog kidney cells (MDCK) after transformation by Harvey murine sarcoma virus. In the present study, we have shown that viral transformation can have differing effects in the ability to modify the production of prostaglandin in cultured cells. For example, the prostaglandin E2 production in rat kidney cells (NRK) is decreased after transformation by Rous sarcoma virus, while production in 3T3 cells is increased markedly after transformation by the same virus. Similarly, SV40 transformation increases prostaglandin E2 production of 3T3 cells and decreases the production in rat thyroid cells (FRTL). These results indicate that the biosynthetic pathway for prostaglandin production has varying susceptibility following viral transformation and the effect of transformation depends more on the type of cell than virus. Taking advantage of the well-defined transforming proteins encoded by polyomavirus, we have further studied the relationship between prostaglandin production in cells and the expression of T antigens in transformed cells. We showed that the expression of middle T antigen, which is associated with a protein kinase and is responsible for phenotype of transformed cells, is required for the change in prostaglandin production in cells. How these changes of prostaglandin production relate to the progression of viral transformation remains to be explored.
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PMID:The effect of viral transformation on prostaglandin production depends on cell type. 302 69

We raised antibodies directed against a synthetic peptide representing an amino acid sequence of the conserved kinase domain of the transforming protein of Fujinami sarcoma virus (FSV) (P140). The antiserum obtained specifically recognized FSV-P140 and its cellular homolog and in addition, it recognized a new cellular protein of 94,000 daltons (NCP94) in avian and mammalian cells. NCP94 was found to be associated with a cyclic nucleotide-independent protein kinase activity that was specific for tyrosine residues. Although NCP94 and FSV-P140 share antigenic determinants, NCP94 is not a cellular homolog of FSV-P140: NCP94 and the previously identified c-fps/fes product were different in their tryptic fingerprints and in their tissue specificities. Thus, the function of NCP94 in normal cells is probably different than that of the c-fps/fes product. NCP94 was expressed in every tissue and cell line that was examined. In chickens, NCP94 levels were highest during embryonic development and NCP94 expression was high in gizzard, brain, and spleen throughout embryonic and adult life. The universal expression of NCP94 suggests that this protein may be involved in an essential function of normal cells. NCP94 may be a new cellular tyrosine kinase of the src gene family.
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PMID:Antipeptide antiserum identifies a widely distributed cellular tyrosine kinase related to but distinct from the c-fps/fes-encoded protein. 302 66

Determination of the mutagenic effects of carcinogenic nickel compounds has been difficult because, like many metals, nickel is poorly or nonmutagenic in procaryotic mutagenicity assays. We attempted to characterize nickel-induced genetic lesions by assessing the effect of nickel chloride on the conditionally defective expression of the v-mos transforming gene in normal rat kidney cells infected with the Murine sarcoma virus mutant ts110 (MuSVts110) retrovirus. MuSVts110 contains an out-of-frame gag gene-mos gene junction that prevents the expression of the v-mos gene at the nonpermissive temperature (39 degrees C). In MuSVts110-infected cells (6m2 cells) grown at 33 degrees C, however, this defect can be suppressed by a splicing event that restores the mos reading frame, allowing the expression of a gag-mos fusion protein which induces the transformed phenotype. The capacity to splice the viral transcript at 33 degrees C, but not at 39 degrees C, is an intrinsic property of the viral RNA. This property allowed us to target the MuSVts110 genome using a positive selection scheme whereby nickel was used to induce genetic changes which resulted in expression of the transformed phenotype at 39 degrees C. We treated 6m2 cells with NiCl2 and isolated foci consisting of cells which had reverted to the transformed phenotype at 39 degrees C. We found that brief nickel treatment increased the reversion frequency of 6m2 cells grown at 39 degrees C sevenfold over the spontaneous reversion frequency. The nickel-induced revertants displayed the following heritable characteristics: They stably maintained the transformed phenotype at 39 degrees C; unlike the MuSVts110 RNA in 6m2 cells, the nickel-induced revertant viral RNA could be spliced efficiently at 39 degrees C; as a consequence of the enhanced accumulation of spliced viral RNA, the nickel-induced revertants produced substantial amounts of the transforming v-mos protein P85gag-mos at 39 degrees C; the nickel-induced revertant P85gag-mos serine kinase, like the parental 6m2 P85gag-mos kinase, was found to be rapidly inactivated at 39 degrees C; however, in the nickel-induced revertants, overproduction of P85gag-mos allowed the transformed state to be maintained; and even though viral RNA processing was much changed, no rearrangements of the viral DNA in the nickel-induced revertant cells were detected by partial restriction analysis.
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PMID:Nickel-induced heritable alterations in retroviral transforming gene expression. 303 2


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