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)

Since the identification of the insulin receptor by insulin-binding activity almost two decades ago, our understanding of the structure and function of the insulin receptor has progressed tremendously. The importance of the intrinsic tyrosine protein kinase activity of the insulin receptor is implied by the fact that the insulin receptor belongs to a family of receptor tyrosine kinases which play a role in growth control, by experiments demonstrating the intimate association of normal kinase activity and insulin action, and by evidence that the intrinsic kinase activity can be regulated under certain conditions. There are still some major gaps in our knowledge concerning the structure/function of the insulin receptor such as how activation of the intrinsic kinase activity of the receptor leads to altered cellular physiology. The kinase may phosphorylate endogenous substrates or autophosphorylation may simply alter beta subunit conformation so it can then interact with an effector system (i.e. a serine kinase) directly, or indirectly through a G-protein. The truth may lie somewhere between these two pathways.
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PMID:Intrinsic kinase activity of the insulin receptor. 215 22

Although CD45 resembles the low Mr protein tyrosine phosphatases (PTPases) from human placenta in its specificity for phosphotyrosyl residues and absolute dependence on sulfhydryl compounds for activity, it also exhibits a number of distinguishing features. Most notably, it displayed substrate specificity in vitro, preferentially dephosphorylating myelin basic protein, over the other substrates tested, with high specific activity. Limited trypsinization of CD45 generated active fragments of approximately 65 kDa that were apparently derived exclusively from the intracellular segment of the molecule. These retained high activity against myelin basic protein, suggesting that this is an intrinsic feature of the PTPase domains and not the result of secondary interactions between the substrate and the putative ligand binding structure. With reduced carboxamidomethylated and maleylated lysozyme as substrate, CD45 was stimulated up to 12-fold by basic compounds such as spermine; divalent metal ions were also stimulatory, most notably Zn2+, which was previously identified as a potent inhibitor of the low Mr PTPases. CD45 was phosphorylated to high stoichiometry by casein kinase-2 (up to 1.5 mol/mol) and also by glycogen synthase kinase 3 (approximately 0.3 mol/mol) and protein kinase C (approximately 0.1 mol/mol); in all cases, no alteration in enzyme activity was detected following these modifications. Autophosphorylated preparations of epidermal growth factor receptor, insulin receptor, and p56lck protein tyrosine kinases were also substrates for CD45 in vitro.
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PMID:CD45, an integral membrane protein tyrosine phosphatase. Characterization of enzyme activity. 216 57

The insulin receptor is a large cell surface glycoprotein that concentrates insulin at the site of action and also initiates responses to insulin. The receptor is a disulfide-linked oligomer comprised of two alpha and two beta subunits. Signal transduction through the insulin receptor appears to require the activation of an intrinsic tyrosine-specific protein kinase activity. A variety of disorders, both acquired and genetic, are associated with the development of insulin resistance and are frequently the result of cellular defects in insulin receptor structure, function, and action. The recent cloning of several mutant receptors from patients with genetic forms of extreme insulin resistance has increased our understanding of insulin resistance on the molecular level.
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PMID:Insulin receptor structure and function in normal and pathological conditions. 218 52

Insulin has both short- and long-term effects on cellular metabolism. The short-term effects are known to involve the insulin receptor, a protein kinase capable of phosphorylating itself and other proteins. The role of the receptor was elucidated by studies of a mutant insulin receptor which lacked kinase activity and inhibited several actions of insulin. The long-term effects of insulin could be demonstrated by its growth-promoting effect on hepatoma cells, and by the suppression in transfected hepatoma cells of hepatitis B virus antigen production in a dose-dependent manner. The process whereby insulin appears to regulate gene expression is not clearly understood.
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PMID:The role of receptor kinase in insulin action and the effects of insulin on human hepatoma cells. 218 55

The effect of protein kinase-C (PKC) inhibition on insulin receptor phosphorylation in HepG2 cells was analyzed by two-dimensional tryptic phosphopeptide maps. In basal cells, there was one major insulin receptor-derived tryptic phosphothreonine peptide and at least four phosphoserine peptides. Phorbol 12,13-dibutyrate (PDBU) stimulated phosphorylation of the phosphothreonine peptide, some of the basal phosphoserine peptides, and at least one phosphoserine peptide that was not detected in the basal state. Staurosporine completely inhibited the PDBU-mediated phosphorylation. Although staurosporine also inhibited basal phosphorylation of the phosphothreonine peptide, down-regulation of PKC did not, suggesting that PKC does not mediate basal insulin receptor phosphorylation. Insulin treatment resulted in the appearance of four phosphotyrosine peptides. It also stimulated the phosphorylation of at least two phosphoserine peptides. One of these may have been a complex of two or more distinct but poorly resolved phosphopeptides, which was seen in basal cells and a component of which seemed to be stimulated by PDBU. However, neither staurosporine nor down-regulation of PKC diminished insulin-stimulated serine phosphorylation of these peptides, indicating that insulin-stimulated receptor serine phosphorylation did not involve PKC activity. The addition of staurosporine to cells that had been incubated with PDBU resulted in the very rapid decay of phosphorylation of the phosphothreonine-containing peptide, indicating that this site of phosphorylation turns over very rapidly, while some of the other phosphoserine-containing peptides, including the major unique site of phosphorylation stimulated by PDBU, turned over more slowly. Thus, the insulin receptor contains several sites of serine/threonine phosphorylation, some of which are substrates for more than one protein kinase. This may permit complex modulation of insulin receptor functions in response to multiple signalling pathways.
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PMID:The effect of protein kinase-C inhibition on insulin receptor phosphorylation. 219 99

Insulin was found to stimulate the serine/threonine kinase activity of the proto-oncogene product Raf-1. This stimulation was observed in HeLa, NIH 3T3, and Chinese hamster ovary cells, all overexpressing the human insulin receptor. In the HeLa cells, 100 pM insulin gave a significant increase in Raf-1 kinase activity, and 100 nM insulin caused a maximal 2-5-fold increase in activity. The increase in activity was detected after 2 min of insulin treatment and peaked after 5 min. In addition to stimulating Raf-1 kinase activity, insulin caused a shift in the electrophoretic mobility of the Raf-1 protein and an increase in the amount of serine phosphorylation of Raf-1. Moreover, a serine/threonine-specific phosphatase, phosphatase 1, but not two tyrosine-specific phosphatases, was found to deactivate the insulin-activated Raf-1 kinase activity. These findings indicate that insulin activates the serine/threonine kinase activity of the Raf-1 proto-oncogene by increasing its content of phosphoserine.
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PMID:Insulin activates the kinase activity of the Raf-1 proto-oncogene by increasing its serine phosphorylation. 219 70

Several growth factors and mitogens have been shown to activate the proto-oncogene product Raf-1 protein kinase in murine fibroblasts, apparently through a direct agonist-stimulated tyrosine phosphorylation of the Raf-1 protein. We investigated the possibility that insulin could also activate the Raf-1 kinase, since its receptor also contains an intrinsic insulin-activated protein tyrosine kinase activity. In several cell lines expressing relatively large numbers of insulin receptors, insulin rapidly stimulated the phosphorylation of immunoreactive Raf-1 protein. In H35 cells, a line of well differentiated rat hepatoma cells, the effect of insulin was maximal by 6 min and at 7 nM insulin and occurred normally in cells virtually completely depleted of protein kinase C activity. The insulin-stimulated increase in Raf-1 protein phosphorylation occurred concurrently with a 3-fold increase in Raf-1 protein kinase activity. However, phosphoamino acid analysis showed that only phosphoserine and a trace of phosphothreonine were present in the Raf-1 protein after insulin stimulation of the cells. This was true even when investigated at shorter times (4 min) after insulin stimulation and despite the use of phosphotyrosine phosphatase inhibitors. We conclude that insulin can rapidly activate the Raf-1 kinase in some insulin-sensitive cell types but that this activation probably occurs through a mechanism distinct from direct phosphorylation of the Raf-1 protein by the insulin receptor protein tyrosine kinase.
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PMID:Insulin activates the Raf-1 protein kinase. 219 71

Previous studies have indicated that turkey erythrocyte and rat liver membranes contain endogenous alpha beta heterodimeric insulin receptors in addition to the disulphide-linked alpha 2 beta 2 heterotetrameric complexes characteristic of most cell types. We utilized 125I-insulin affinity cross-linking to examine the structural properties of insulin receptors from rat liver and turkey erythrocyte membranes prepared in the absence and presence of sulphydryl alkylating agents. Rat liver membranes prepared in the absence of sulphydryl alkylating agents displayed specific labelling of Mr 400,000 and 200,000 bands, corresponding to the alpha 2 beta 2 heterotetrameric and alpha beta heterodimeric insulin receptor complexes respectively. In contrast, affinity cross-linking of membranes prepared with iodoacetamide (IAN) or N-ethylmaleimide identified predominantly the alpha 2 beta 2 heterotetrameric insulin receptor complex. Similarly, affinity cross-linking and solubilization of intact turkey erythrocytes in the presence of IAN resulted in exclusive labelling of the alpha 2 beta 2 heterotetrameric insulin receptor complex, whereas in the absence of IAN both alpha 2 beta 2 and alpha beta species were observed. Turkey erythrocyte alpha 2 beta 2 heterotetrameric insulin receptors from IAN-protected membranes displayed a 3-4-fold stimulation of beta subunit autophosphorylation and substrate phosphorylation by insulin, equivalent to that observed in intact human placenta insulin receptors. Turkey erythrocyte alpha beta heterodimeric insulin receptors, prepared by defined pH/dithiothreitol treatment of IAN-protected membranes, were also fully competent in insulin-stimulated protein kinase activity compared with alpha beta heterodimeric human placenta receptors. In contrast, endogenous turkey erythrocyte alpha beta heterodimeric insulin receptors displayed basal protein kinase activity which was insulin-insensitive. These data indicate that native turkey erythrocyte and rat liver insulin receptors are structurally and functionally similar to alpha 2 beta 2 heterotetrameric human placenta insulin receptors. The alpha beta heterodimeric insulin receptors previously identified in these tissues most likely resulted from disulphide bond reduction and denaturation of the alpha 2 beta 2 holoreceptor complexes during membrane preparation.
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PMID:The endogenous functional turkey erythrocyte and rat liver insulin receptor is an alpha 2 beta 2 heterotetrameric complex. 222 23

Phosphorylation of the insulin receptor beta-subunit on serine/threonine residues by protein kinase C reduces both receptor kinase activity and insulin action in cultured cells. Whether this mechanism regulates insulin action in intact animals was investigated in rats rendered insulin-resistant by 3 days of starvation. Insulin-stimulated autophosphorylation of the partially purified hepatic insulin receptor beta-subunit was decreased by 45% in starved animals compared to fed controls. This autophosphorylation defect was entirely reversed by removal of pre-existing phosphate from the receptor with alkaline phosphatase, suggesting that increased basal phosphorylation on serine/threonine residues may cause the decreased receptor tyrosine kinase activity. Tryptic removal of a C-terminal region of the receptor beta-subunit containing the Ser/Thr phosphorylation sites similarly normalized receptor autophosphorylation. To investigate which kinase(s) may be responsible for such increased Ser/Thr phosphorylation in vivo, protein kinase C and cAMP-dependent protein kinase A in liver were studied. A 2-fold increase in protein kinase C activity was found in both cytosol and membrane extracts from starved rats as compared to controls, while protein kinase A activity was diminished in the cytosol of starved rats. A parallel increase in protein kinase C was demonstrated by immunoblotting with a polyclonal antibody which recognizes several protein kinase C isoforms. These findings suggest that in starved, insulin-resistant animals, an increase in hepatic protein kinase C activity is associated with increased Ser/Thr phosphorylation which in turn decreases autophosphorylation and function of the insulin receptor kinase.
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PMID:Increased protein kinase C activity is linked to reduced insulin receptor autophosphorylation in liver of starved rats. 235 98

Phosphotyrosine-containing proteins are minor components of normal cells which appear to be associated primarily with the regulation of cellular metabolism and growth. The insulin receptor is a tyrosine-specific protein kinase, and one of the earliest detectable responses to insulin binding is activation of this kinase and autophosphorylation of its beta-subunit. Tyrosine autophosphorylation activates the phosphotransferase in the beta-subunit and increases its reactivity toward tyrosine phosphorylation of other substrates. When incubated in vitro with [gamma-32P]ATP and insulin, the purified insulin receptor phosphorylates various proteins on their tyrosine residues. However, so far no proteins other than the insulin receptor have been identified as undergoing tyrosine phosphorylation in response to insulin in an intact cell. Here, using anti-phosphotyrosine antibodies, we have identified a novel phosphotyrosine-containing protein of relative molecular mass (Mr) 185,000 (pp185) which appears during the initial response of hepatoma cells to insulin binding. In contrast to the insulin receptor, pp185 does not adhere to wheat-germ agglutininagarose or bind to anti-insulin receptor antibodies. Phosphorylation of pp185 is maximal within seconds after exposure of the cells to insulin and exhibits a dose-response curve similar to that of receptor autophosphorylation, suggesting that this protein represents the endogenous substrate for the insulin receptor kinase.
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PMID:Insulin rapidly stimulates tyrosine phosphorylation of a Mr-185,000 protein in intact cells. 241 72


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