Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.10.1 (ERK)
 95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human platelets contain platelet-derived growth factor (PDGF) in their alpha-granules which is released during platelet exocytosis. We show by immunoprecipitation and 125I-PDGF binding experiments that human platelets have functionally active PDGF alpha-receptors, but not beta-receptors. The PDGF alpha-receptor (PDGFR-alpha) was identified as a 170-kDa glycosylated protein-tyrosine kinase as found in other cell types. Stimulation of platelets with 0.1 unit/ml thrombin resulted in a significant increase (2-5-fold) of the tyrosine phosphorylation of the PDGFR-alpha, as determined by immunoprecipitation with phosphotyrosine antiserum as well as with PDGFR-alpha antiserum. The observed thrombin-induced autophosphorylation of the PDGFR-alpha was inhibited by the addition of a neutralizing monoclonal PDGF antibody. Thus, our results suggest that the platelet PDGFR-alpha is stimulated in an autocrine manner by PDGF secreted during platelet activation. Preincubation of platelets with PDGF inhibited thrombin-induced platelet aggregation and secretion of ATP + ADP and beta-hexosaminidase. Thrombin-induced platelet aggregation was also reversed when PDGF was added 30 s after thrombin stimulation. Inhibition of the autocrine PDGF pathway during platelet activation by the PDGF antibody led to a potentiation of thrombin-induced beta-hexosaminidase secretion. Thus, the PDGFR-alpha takes part in a negative feedback regulation during platelet activation. Our demonstration of PDGF alpha-receptors on human platelets and its inhibitory function during platelet activation identifies a new possible role of PDGF in the regulation of thrombosis.
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PMID:Negative feedback regulation of human platelets via autocrine activation of the platelet-derived growth factor alpha-receptor. 818 64

In this report we present some of the biochemical properties of the enzyme, here called pp28(PTK), isolated from particulate fraction of rat spleen (1). The kinase is very susceptible for polyions as regulators of the enzymatic activity. The polyanions like dextran sulfate or heparin inhibited, and polycations such as spermidin, protamin, poly-L-lysine and some random polypeptides containing tyrosine besides a basic amino acid, stimulated the enzyme markedly. The kinase showed high sensitivity towards class IA salts. In the casein phosphorylation reaction the apparent Km value for ATP was 4 microM. An unusual property is associated with autophosphorylation which leads to a reduced activity towards external substrates. Some kinase inhibitors described in the literature were tested for their potency.
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PMID:Biochemical properties of a novel 28KDA protein tyrosine kinase partially purified from the particulate fraction of rat spleen. 826 1

G protein-coupled receptor kinases (GRKs) such as rhodopsin kinase and the beta-adrenergic receptor kinase (beta ARK) play an important role in agonist-specific phosphorylation and desensitization of G protein-coupled receptors. GRK5 is a recently identified member of the GRK family that has greater homology with rhodopsin kinase than with beta ARK. To further characterize the activity of GRK5, it has been overexpressed in Sf9 insect cells and purified by successive chromatography on S-Sepharose and Mono S columns. GRK5 phosphorylates the beta 2-adrenergic receptor (beta 2AR), m2 muscarinic cholinergic receptor, and rhodopsin in an agonist-dependent manner to maximal stoichiometries of approximately 2.5, 1.5, and 1 mol of phosphate/mol of receptor, respectively, with Km values of approximately 0.5 microM for the beta 2AR, approximately 16 microM for rhodopsin, and approximately 24 microM for ATP. Peptide phosphorylation studies suggest that in contrast to beta ARK and rhodopsin kinase, GRK5 preferentially phosphorylates on nonacidic peptides with a Km of approximately 1.5 mM. Heparin and dextran sulfate were found to be potent inhibitors of GRK5 with IC50 values of approximately 1 nM, thereby being at least 150-fold more potent on GRK5 than on beta ARK. GRK5 can also be activated by polycations, with 10 microM polylysine promoting an approximately 2.6-fold activation. Overall, these studies demonstrate that GRK5 has unique properties that distinguish it from other members of the GRK family and that likely play an important role in modulating its mechanism of action.
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PMID:Expression, purification, and characterization of the G protein-coupled receptor kinase GRK5. 828 67

The efficiency of a preservation medium, histidine-buffered lactobionate solution (HBLS), was determined by measuring post-ischemic recoveries of ATP and intracellular pH under Krebs-Henseleit buffer (KHB) perfusion. We used NMR spectroscopy to study the effect of 24-h cold ischemia, followed by 4 degrees C then 37 degrees C reperfusion on the isolated rat liver. Three media were compared: University of Wisconsin solution (UW-lactobionate); Bretschneider's solution (HTK); HBLS and HBLS supplemented with 2 mM Gly and 2 mM Cys (HBLSg2) or with 10 mM Gly and 2 mM Cys (HBLSg10). All values were compared to control values measured during pre-ischemic cold perfusion with KHB (ATP = 8.60 +/- 0.6 mumol/g of dry weigh and pH(in) = 7.41 +/- 0.05). The main result from 31P NMR data concerned ATP recovery during cold reperfusion, which was significantly higher in the HBLS group (112 +/- 10%) as compared to the UW and HTK groups (around 66%). The presence of glycine decreased ATP recovery (88 +/- 8% in HBLSg2, 79 +/- 15% in HBLSg10). Higher values of recovered pHin were observed in livers stored in histidine buffered solutions (around 7.30) as compared to UW (around 7.20); histidine was by 13C NMR proved to accumulate in the liver cells, thus ensuring a good buffering capacity. The thermal transition induced a decrease in both ATP level and pHin in all groups. This might be the result of a stimulation of the carbohydrate metabolism (as demonstrated by 13C NMR) especially when glycine was present in the storage solution.
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PMID:31P NMR studies of rat liver cold preservation with histidine-buffered lactobionate solution. 830 4

This study investigates how far mitochondrial swelling in the ischemic heart is influenced by factors pertaining to anaerobic energy turnover. Canine hearts were arrested by aortic cross clamping or cardioplegically with St. Thomas or HTK solution and incubated at 25 degrees C in the solution used for cardiac arrest. Samples of the left ventricle were taken at the end of cardiac arrest and during ischemia for structural evaluation and biochemical analysis. The extracellular pH in the interventricular septum was measured. Mitochondrial swelling was determined with the surface to volume ratio, a parameter independent of the reference space. Values obtained for different swelling were related to defined metabolite concentrations and pHe values to establish possible correlations between structural and biochemical parameters in the ischemic heart. At the onset of ischemia and during the breakdown of creatine phosphate (CP) to 3 mumol/g wet weight mitochondrial volume depends on the method of cardiac arrest and does not increase significantly in any of the three groups. The degree of mitochondrial swelling after depletion of CP correlates with the decline in ATP independent of the form of cardiac arrest. Characteristic values of the surface to volume ratio ascertained at different times of ischemia for all groups correspond to determined ATP concentrations. Acid pHe values seem to intensify mitochondrial swelling. With increased lactate concentrations mitochondria swell, but first initially the degree of swelling differs significantly in the forms of cardiac arrest investigated. Thus, the surface to volume ratio is a powerful and valid ultrastructural parameter, which makes correlations between mitochondrial structure and metabolism possible and furthermore indicates a strong correlation between mitochondrial swelling and ATP-concentration in the ischemic heart.
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PMID:Close correlations between mitochondrial swelling and ATP-content in the ischemic canine myocardium. A combined morphometric and biochemical study. 833 76

The response of the myocardium to prolonged or chronic ischemia may differ from the well documented changes that occur acutely subsequent to the onset of hypoperfusion. Therefore, we have examined in an instrumented canine model and using spatially localized spectroscopy to achieve transmural differentiation, the myocardial HEP and Pi levels as well as wall thickening in situ during prolonged ischemia induced by sustained coronary artery stenosis. The results demonstrate that subtotal coronary artery occlusion causes immediate and transmurally inhomogeneous decreases in the myocardial HEP content and increase in the Pi/CP ratio; however, during prolonged mild hypoperfusion, metabolic changes occur which lead to statistically significant recovery of CP (but not ATP) and disappearance of Pi despite the persistence of reduced blood flow and oxygen supply. Upon release of the occlusion, the previously ischemic muscle recovered blood flow, and some (but not all) of its preischemic contractile function without parallel changes in the HEP levels. It is concluded that normal HEP and Pi levels cannot be equated with either the absence of underperfusion or insensitivity of NMR spectroscopy to ischemia. Rather, it is imperative that both functional and spectroscopic measurements are performed simultaneously to distinguish between ischemic myocardium which is adapted versus unadapted to the hypoperfusion.
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PMID:Responses of myocardial high energy phosphates and wall thickening to prolonged regional hypoperfusion induced by subtotal coronary stenosis. 837 71

A cDNA encoding the human homologue of mouse RYK (related to receptor tyrosine kinases) has been cloned from an interleukin 1 (IL-1)-stimulated human hepatoma cDNA library by cross-species hybridization using the mouse RYK cDNA as a probe. The sequence of the 3067-bp cDNA clone encoding human RYK predicts a transmembrane protein with a cytoplasmic domain that contains the consensus sequences (subdomains I-XI) of the protein tyrosine kinase (PTK) family. The highly conserved motif -D-F-G- (subdomain VII) of the catalytic domain of other receptor-type tyrosine kinases is altered to -D-N-A- in human RYK. In addition, a number of other changes were found in the ATP binding site (subdomains I and II) and the motif [-I-H-R-D-L-A-A-R-N-] found in subdomain VI. Comparison of the human and mouse RYK sequences shows a 92% conservation at the nucleotide level and 97% at the amino acid level. There was no significant homology between the extracellular domain of RYK and the other families of receptor tyrosine kinases described to date. RYK therefore appears to define a new subclass of receptor-type tyrosine kinases whose structure has remained highly conserved across species.
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PMID:Molecular cloning and chromosomal localisation of the human homologue of a receptor related to tyrosine kinases (RYK). 838 29

Activation of the cyclin-dependent protein kinases p34cdc2 and p33cdk2 requires binding with a cyclin partner and phosphorylation on the first threonine residue in the sequence THEVVTLWYRAPE. We present evidence that this threonine residue, number 160 in p33cdk2, can be specifically phosphorylated by a cdc2-related protein kinase from Xenopus oocytes called p40MO15. Binding to cyclin A and phosphorylation of this threonine are both required to activate fully the histone H1 kinase activity of p33cdk2. In cell extracts, a portion of p40MO15 is found in a high molecular weight complex that is considerably more active than a lower molecular weight form. Wild-type MO15 protein expressed in bacteria does not possess kinase activity, but acquires p33cdk2-T160 kinase activity after incubation with cell extract and ATP. We conclude that p40MO15 corresponds to CAK (cdc2/cdk2 activating kinase) and speculate that, like p33cdk2 and p34cdc2, p40MO15 requires activation by phosphorylation and association with a companion subunit.
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PMID:The cdc2-related protein p40MO15 is the catalytic subunit of a protein kinase that can activate p33cdk2 and p34cdc2. 839 83

While screening a chicken kidney cDNA library for the normal homolog of the yes oncogene, we isolated a clone that encodes a novel non-receptor type protein tyrosine kinase of the Src family. We named this gene product Yrk (York), as an acronym for Yes-related kinase. As predicted from the cDNA sequence, the Yrk protein consists of 536 amino acids and has all the canonical features of a Src kinase. At the amino terminus it contains a myristylation signal, followed by a unique domain, SH3 and SH2 motifs, an ATP binding site, a kinase region and a carboxy-terminal sequence with a potential regulatory tyrosine at position 530. The sequence of the Yrk protein showed 79% identity with human Fyn and 72% identity with chicken Yes. To eliminate the possibility that the Yrk protein is an avian homolog of mammalian Fyn, we isolated and sequenced the chicken fyn cDNA. The sequence data together with Southern and Northern blot analyses showed that the chicken yrk gene is distinct from the chicken fyn gene. Antibodies generated against the unique domain of the yrk protein expressed in bacteria precipitated a 60-kDa protein that was active in an immune complex kinase assay and was phosphorylated on tyrosine. Expression of the Yrk protein in adult chicken tissues was elevated in cerebellum and spleen. Relatively high levels of Yrk were also found in lung and skin.
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PMID:A novel Yes-related kinase, Yrk, is expressed at elevated levels in neural and hematopoietic tissues. 845 40

The beta-adrenergic receptor kinase mediates agonist-dependent phosphorylation of beta-adrenergic receptors, which is thought to represent the first step of homologous desensitization. We have expressed bovine and human beta ARK1 in Sf9 cells and purified them to apparent homogeneity in milligram quantities. The Km-values of the enzyme were 3.8 microM for rhodopsin and 22 microM for ATP; the Vmax-value was 9.9 mol phosphate/mol beta ARK/min. These data indicate that the two recombinant kinases were at least as active as preparations previously obtained from bovine brain. There were no differences in the functional activity of human and bovine beta ARK.
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PMID:Purification and functional characterization of beta-adrenergic receptor kinase expressed in insect cells. 850 60


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