Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: DrugBank:EXPT02079 (lysine)
 58,762 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A 'MAP kinase activator' was purified several thousand-fold from insulin-stimulated rabbit skeletal muscle, which resembled the 'activator' from nerve growth factor-stimulated PC12 cells in that it could be inactivated by incubation with protein phosphatase 2A, but not by protein tyrosine phosphatases and its apparent molecular mass was 45-50 kDa. In the presence of MgATP, 'MAP kinase activator' converted the normal 'wild-type' 42 kDa MAP kinase from an inactive dephosphorylated form to the fully active diphosphorylated species. Phosphorylation occurred on the same threonine and tyrosine residues which are phosphorylated in vivo in response to growth factors or phorbol esters. A mutant MAP kinase produced by changing a lysine at the active centre to arginine was phosphorylated in an identical manner by the 'MAP kinase activator', but no activity was generated. The results demonstrate that 'MAP kinase activator' is a protein kinase (MAP kinase kinase) and not a protein that stimulates the autophosphorylation of MAP kinase. MAP kinase kinase is the first established example of a protein kinase that can phosphorylate an exogenous protein on threonine as well as tyrosine residues.
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PMID:MAP kinase activator from insulin-stimulated skeletal muscle is a protein threonine/tyrosine kinase. 131 93

An hepatic protein kinase that phosphorylates microtubule-associated protein 2 (MAP-2) on Ser/Thr residues is markedly activated after intraperitoneal injection of cycloheximide in the rat. The enzyme has been purified greater than 10,000-fold to near homogeneity and corresponds to a 54-kDa polypeptide, based on auto-phosphorylation, renaturation of activity from sodium dodecyl sulfate gels, and gel filtration. The protein kinase activity is unaffected by prior autophosphorylation, Ca2+, diacylglycerol and phospholipids, cyclic nucleotides, staurosporine, and protein kinase inhibitor, but can be totally and specifically deactivated by the Ser/Thr protein phosphatase 2A. The enzyme is inhibited completely but reversible by transition metals and p-chloromercuribenzoate, and is strongly stimulated by poly-L-lysine toward most, but not all protein substrates. The activity of the cycloheximide-stimulated MAP-2 kinase (pp54 MAP-2 kinase) toward potential polypeptide substrates was compared to that of an insulin-stimulated MAP-2 kinase (pp42 MAP-2 kinase). Although both MAP-2 kinases exhibited little or no ability to phosphorylate histones and casein, the two kinases had a distinguishable substrate specificity. At comparable MAP-2 phosphorylating activities, pp42 MAP-2 kinase, but not pp54 MAP-2 kinase, phosphorylated and activated the Xenopus S6 protein kinase II. Moreover, pp42 MAP-2 kinase phosphorylated myelin basic protein at 10-12-fold higher rates than did pp54 MAP-2 kinase. Cycloheximide-activated pp54 MAP-2 protein kinase appears to be a previously uncharacterized protein kinase that is itself regulated through Ser/Thr phosphorylation and, perhaps, polypeptide regulators with basic domains. The identity of the upstream regulatory elements and the native substrates remain to be established.
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PMID:pp54 microtubule-associated protein 2 kinase. A novel serine/threonine protein kinase regulated by phosphorylation and stimulated by poly-L-lysine. 217 Mar 74

The Ste20p protein kinase was immunopurified from yeast cells and analyzed in an in vitro assay system. Ste20p immune complexes exhibited autophosphorylating activity at serine and threonine residues and specifically phosphorylated a bacterially expressed glutathione S-transferase (GST) fusion of Ste11p (a mitogen-activated protein or extracellular signal-regulated kinase kinase (MEK) kinase homologue) at serine and threonine residues. In contrast, GST fusions either of Ste7p (a MEK homologue) or the beta-subunit of the mating response G-protein and immunoprecipitated Ste5p were not phosphorylated by the Ste20p immune complexes. Myelin basic protein was identified as an excellent in vitro substrate, whereas histone H1 was only poorly phosphorylated. Evidence was obtained that autophosphorylation might play a regulatory role for the in vitro kinase activity. The in vitro activity was found to be Ca(2+)-independent. Both the in vivo and in vitro activities were abolished by mutational changes of either the conserved lysine residue 649 within the ATP binding site or threonine 777 between the catalytic subdomains VII and VIII. Wild-type Ste20p and the catalytically inactive T777A mutant were identified as phosphoproteins in vivo. The phosphorylation occurred at serine and threonine residues independent of pheromone stimulation. Based on the genetically determined significance of Ste20p in pheromone signal transduction and on our in vitro studies, we propose the model that Ste20p represents a yeast MEK kinase kinase whose function is to link G-protein-coupled receptors through G beta gamma to a mitogen-activated protein kinase module.
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PMID:Molecular characterization of Ste20p, a potential mitogen-activated protein or extracellular signal-regulated kinase kinase (MEK) kinase kinase from Saccharomyces cerevisiae. 760 57

Integrins can function as signal-transducing receptors capable of modulating cell growth and gene expression (Juliano, R. L., and Haskill, S. (1993) J. Cell Biol. 120, 577-585; Hynes, R. O. (1992) Cell 69, 11-25). An early event in integrin signaling in fibroblasts and other cells involves activation of pp125FAK, a cytoplasmic tyrosine kinase (Hanks, S. K., Calalb, M. B., Harper, M. C., and Patel, S. K. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 8487-8491; Schaller, M. D., Borgman, C. A., Cobb, B. S., Vines, R. R., Reynolds, A. B., and Parsons, J. T. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 5192-5196). Here we report a novel aspect of integrin-mediated signal transduction. We demonstrate that adhesion of cells to substrata coated with extracellular matrix proteins, or with a synthetic peptide containing the RGD sequence, can cause activation of mitogen-activated protein (MAP) kinases in 3T3 or REF52 fibroblasts. Activation of MAP kinases seems to depend on integrin engagement rather than simply on cell attachment. Thus, MAP kinases are activated when cells adhere to substrata coated with the integrin ligands fibronectin or laminin, but not when cells adhere to poly-D-lysine, a nonspecific adhesion-promoting polypeptide. Treatment of cells with cytochalasin D, an inhibitor of actin microfilament assembly, almost completely blocks adhesion-induced MAP kinase activation, indicating a critical role for the cytoskeleton. In REF52 cells, we have observed that activation of MAP kinases is accompanied by redistribution of the protein to the nucleus, suggesting that the activated kinases may impinge on factors regulating gene expression. Thus, integrin-mediated cell adhesion seems a sufficient stimulus to cause activation and nuclear translocation of MAP kinases. This may have important implications for the regulation of cell growth and differentiation by the extracellular matrix.
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PMID:Integrin-mediated cell adhesion activates mitogen-activated protein kinases. 792 88

The substrate specificity of mitogen-activated protein (MAP) kinase-activated protein kinase-2 (MAPKAP kinase-2) was investigated by using synthetic peptides related to the N-terminus of glycogen synthase. The minimum sequence required for efficient phosphorylation was found to be Xaa-Xaa-Hyd-Xaa-Arg-Xaa-Xaa-Ser-Xaa-Xaa, where Hyd is a bulky hydrophobic residue (Phe > Leu > Val >> Ala), and the peptide Lys-Lys-Phe-Asn-Arg-Thr-Leu-Ser-Val-Ala was phosphorylated with a Km of 9.3 microM and Vmax. of 10 mumol/min per mg. MAPKAP kinase-1 (a homologue of ribosomal protein S6 kinase) also requires an arginine three residues N-terminal to the serine (position n-3), but not a hydrophobic residue at position n-5. Neither MAPKAP kinase-1 nor MAPKAP kinase-2 could tolerate a proline residue at position n + 1, indicating that their specificities do not overlap with that of MAP kinase. The specificity of calmodulin-dependent protein kinase-II resembled that of MAPKAP kinase-2, except that it could tolerate replacement of the arginine by a lysine and the phosphorylation-site serine by a threonine residue. Partial cDNAs encoding MAPKAP kinase-2 were isolated from rabbit and human skeletal muscle and human teratocarcinoma libraries, and Northern-blotting experiments revealed a single 3.3 kb mRNA transcript present at similar levels in six human tissues examined. The catalytic domain was most similar (35-40% identity) to calmodulin-dependent protein kinases II and IV, phosphorylase kinase, putative serine kinase H1 and the C-terminal domain of MAPKAP kinase-1, which form one branch of the protein kinase phylogenetic tree. The sequence N-terminal to the catalytic domain is proline-rich and contains two putative SH3-binding sites. The threonine residue phosphorylated by MAP kinase lies immediately C-terminal to the catalytic domain and is followed by a nuclear localization signal, Lys-Lys-(Xaa)10-Lys-Arg-Arg-Lys-Lys, near the C-terminus.
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PMID:The substrate specificity and structure of mitogen-activated protein (MAP) kinase-activated protein kinase-2. 828 84

The Ras guanylnucleotide exchange protein SOS undergoes feedback phosphorylation and dissociation from Grb2 following insulin receptor kinase activation of Ras. To determine the serine/threonine kinase(s) responsible for SOS phosphorylation in vivo, we assessed the role of mitogen-activated, extracellular-signal-regulated protein kinase kinase (MEK), extracellular-signal-regulated protein kinase (ERK), and the c-JUN protein kinase (JNK) in this phosphorylation event. Expression of a dominant-interfering MEK mutant, in which lysine 97 was replaced with arginine (MEK/K97R), resulted in an inhibition of insulin-stimulated SOS and ERK phosphorylation, whereas expression of a constitutively active MEK mutant, in which serines 218 and 222 were replaced with glutamic acid (MEK/EE), induced basal phosphorylation of both SOS and ERK. Although expression of the mitogen-activated protein kinase-specific phosphatase (MKP-1) completely inhibited the insulin stimulation of ERK activity both in vitro and in vivo, SOS phosphorylation and the dissociation of the Grb2-SOS complex were unaffected. In addition, insulin did not activate the related protein kinase JNK, demonstrating the specificity of insulin for the ERK pathway. The insulin-stimulated and MKP-1-insensitive SOS-phosphorylating activity was reconstituted in whole-cell extracts and did not bind to a MonoQ anion-exchange column. In contrast, ERK1/2 protein was retained by the MonoQ column, eluted with approximately 200 mM NaCl, and was MKP-1 sensitive. Although MEK also does not bind to MonoQ, immunodepletion analysis demonstrated that MEK is not the insulin-stimulated SOS-phosphorylating activity. Together, these data demonstrate that at least one of the kinases responsible for SOS phosphorylation and functional dissociation of the Grb2-SOS complex is an ERK-independent but MEK-dependent insulin-stimulated protein kinase.
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PMID:Insulin stimulation of a MEK-dependent but ERK-independent SOS protein kinase. 855 85

Phosphohistidine goes undetected in conventional studies of protein phosphorylation, although it may account for 6% of total protein phosphorylation in eukaryotes. Procedures for studying protein N- kinases are described. Genes whose products are putative protein histidine kinases occur in a yeast and a plant. In rat liver plasma membranes, activation of the small G-protein, Ras, causes protein histidine phosphorylation. Cellular phosphatases dephosphorylate phosphohistidine. One eukaryotic protein histidine kinase has been purified, and specific proteins phosphorylated on histidine have been observed. There is a protein arginine kinase in mouse and protein lysine kinases in rat. Protein phosphohistidine may regulate the mitogen-activated protein kinase cascade.
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PMID:Protein kinases and phosphatases that act on histidine, lysine, or arginine residues in eukaryotic proteins: a possible regulator of the mitogen-activated protein kinase cascade. 857 21

Among the protein kinases, an absolutely conserved lysine in subdomain II is required for high catalytic activity. This lysine is known to interact with the substrate ATP, but otherwise its role is not well understood. We have used biochemical and structural methods to investigate the function of this lysine (K52) in phosphoryl transfer reactions catalyzed by the MAP kinase ERK2. The kinetic properties of activated wild-type ERK2 and K52 mutants were examined using the oncoprotein TAL2, myelin basic protein, and a designed synthetic peptide as substrates. The catalytic activities of K52R and K52A ERK2 were lower than that of wild-type ERK2, primarily as a consequence of reductions in kcat. Further, there was little difference in Km for ATP, but the Km,app for peptide substrate was higher for the K52 mutants. The three-dimensional structure of unphosphorylated K52R ERK2 in the absence and presence of bound ATP was determined and compared with the structure of unphosphorylated wild-type ERK2. ATP adopted a well-defined but distinct binding mode in K52R ERK2 compared to the binding mode in the wild-type enzyme. The structural and kinetic data show that mutation of K52 created a nonproductive binding mode for ATP and suggest that K52 is essential for orienting ATP for catalysis.
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PMID:Mutation of position 52 in ERK2 creates a nonproductive binding mode for adenosine 5'-triphosphate. 863 22

The PDGF beta-receptor in which the active-site lysine in the kinase domain has been mutated to arginine (K634R) tacks intrinsic kinase activity. When expressed in HepG2 cells, the kinase-inactive PDGF beta-receptor was tyrosine phosphorylated in response to PDGF-BB. Previously, HepG2 cells were thought to be devoid of PDGF alpha-receptor primarily due to lack of specific antibody which precluded detection of the PDGF alpha-receptor. In fact, these cells express low levels of PDGF alpha-receptor. In HepG2 cells that express the kinase-inactive PDGF beta-receptor, PDGF-BB activates the PDGF alpha-receptors to trans phosphorylate the kinase-inactive PDGF beta-receptor in an intermolecular fashion. As a result, stimulation of HepG2 cells that express the kinase-inactive receptor leads to activation of serine/threonine kinases of the MAP kinase cascade which include RAF-1, MEK-1 and p42 MAP kinase. In contrast, the kinase-inactive receptor does not activate any signaling pathways when it is expressed in PC12 cells which do not express the endogenous PDGF alpha-receptor. Thus, the kinase-inactive K634R PDGF beta-receptor is able to enhance PDGF-BB signaling in HepG2 cells that express the PDGF alpha-receptor.
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PMID:The kinase-inactive PDGF beta-receptor mediates activation of the MAP kinase cascade via the endogenous PDGF alpha-receptor in HepG2 cells. 870 May 41

Fluid shear stress modulates vascular function and structure by stimulating mechanosensitive endothelial cell signal events. Cell adhesion, mediated by integrin-matrix interactions, also regulates intracellular signaling by mechanosensitive events. To gain insight into the role of integrin-matrix interactions, we compared tyrosine phosphorylation and extracellular signal-regulated kinase (ERK1/2) activation in adhesion- and shear stress-stimulated human umbilical vein endothelial cells (HUVEC). Adhesion of HUVEC to fibronectin, but not to poly-L-lysine, rapidly activated ERK1/2. Fluid shear stress (12 dyn/cm2) enhanced ERK1/2 activation stimulated by adhesion, suggesting the presence of a separate pathway. Two differences in signal transduction were identified: focal adhesion kinase phosphorylation was increased rapidly by adhesion but not by shear stress; and ERK1/2 activation in response to adhesion was inhibited to a significantly greater extent when actin filaments were disrupted by cytochalasin D. Two similarities in activation of ERK1/2 were observed: protein kinase C (PKC) activity was necessary as shown by complete inhibition when PKC was downregulated; and an herbimycin-sensitive (genistein- and tyrphostin-insensitive) tyrosine kinase was required. c-Src was identified as a candidate tyrosine kinase as it was activated by both shear stress and adhesion. These findings suggest that adhesion and shear stress activate ERK1/2 via a shared pathway that involves an herbimycin-sensitive tyrosine kinase and PKC. In addition, shear stress activates ERK1/2 through another pathway that is partially independent of cytoskeletal integrity.
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PMID:Mitogen-activated protein kinase (ERK1/2) activation by shear stress and adhesion in endothelial cells. Essential role for a herbimycin-sensitive kinase. 895 27


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