Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.26 (GSK)
 6,788 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have isolated the full-length sequence for a unique human kinase, designated TTK. TTK was initially identified by screening of a T cell expression library with anti-phosphotyrosine antibodies. The kinases most closely related to TTK are the SPK1 serine, threonine and tyrosine kinase, the Pim1, PBS2, and CDC2 serine/threonine kinases, and the TIK kinase which was also identified through screening of an expression library with anti-phosphotyrosine antibodies. However, the relationships are distant with less than 25% identity. Nevertheless, TTK is highly conserved throughout phylogeny with hybridizing sequences being detected in mammals, fish, and yeast. TTK mRNA is present at relatively high levels in testis and thymus, tissues which contain a large number of proliferating cells, but is not detected in most other benign tissues. Freshly isolated cells from most malignant tumors assessed expressed TTK mRNA. As well, all rapidly proliferating cell lines tested expressed TTK mRNA. Escherichia coli expressing the complete kinase domain of TTK contain markedly elevated levels of phosphoserine and phosphothreonine as well as slightly increased levels of phosphotyrosine. Taken together, these findings suggest that expression of TTK, a previously unidentified member of the family of kinases which can phosphorylate serine, threonine, and tyrosine hydroxyamino acids, is associated with cell proliferation.
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PMID:Expression of TTK, a novel human protein kinase, is associated with cell proliferation. 163 25

Washing buffy-coat free erythrocyte concentrates three times in bottles used for blood storage will diminish their leukocyte content to 0.22 +/- 0.11 x 10(9) per TE (= 9% of the initial value in whole blood, and the thrombocyte content to 0.3 +/- 0.5 x 10(9) per day (= 2% of the initial value in whole blood). Even 50% of leukocytes (mainly lymphocytes) and 80% of thrombocytes are eliminated simply by buffy coat separation. 30% of erythrocytes are lost by the washing process. Due to increasing haemolysis (0.22%) a subsequent storage of 24 hours should not be exceeded for washed erythrocyte concentrates. Further quality parameters, such as morphological index, pH, ATP, 2,3-P2G and K+ and Na+, were investigated. As far as selected quality parameters are concerned, washing erythrocyte concentrates three times in bottles for blood storage may be compared with washing them once in blood bags. The present findings confirm the conclusion that the washing of erythrocyte concentrates with a solution of sodium chloride in order to eliminate leukocytes may for the most part exclude non-haemolytic febrile transfusion reactions, but not immunization. More effective procedures of eliminating leukocytes, such as filtration, TTK or even glycerin, treatment of erythrocyte concentrates without cryoconservation, are indispensable.
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PMID:[Improving the quality of washed and buffy coat-free erythrocyte concentrates]. 169 88

TPK-IIB, a spleen tyrosine protein kinase devoid of autophosphorylation activity (Brunati, A. M., and Pinna, L. A. (1988) Eur. J. Biochem. 172, 451-457), has been purified to near homogeneity and assayed for its ability to phosphorylate the synthetic peptides EDNEYTA and EPQYQPA reproducing the two conserved phosphoacceptor sites of pp60c-src (Tyr-416 and Tyr-527). While EPQYQPA was phosphorylated with low efficiency (Km = 16.7 mM, Kcat = 14.4), EDNEYTA is an excellent substrate displaying a Km value of 58 microM and a Kcat value of 31.2. The single substitution, in the latter peptide, of the glutamic acid adjacent to the tyrosine by alanine to give EDNAYTA caused a 6-fold increase in the Km. The positive influence on the phosphorylation of the acidic residues at -3 and -4 relative to the tyrosine is indicated by comparison of the kinetic constants for peptides EDAAYAA (Kcat = 4.6, Km 0.325 mM) and QNAAYAA (Kcat 2.4, Km 1.7 mM). Furthermore, when residues in the peptide NEYTA were replaced by alanine, the phosphorylation of the peptides NAYTA and AAYAA, was almost negligible (in terms of Kcat/Km ratio). However, AEYTA, NEYAA and AEYAA were still phosphorylated, albeit less efficiently than NEYTA. The probability that these peptides will adopt a beta-turn is EDNAYTA = EDNEYTA, NAYTA greater than NEYTA, and no predicted beta-turn for AEYTA, NEYAA, and AEYAA. Therefore these results support the concept that an amino-terminal acidic residue(s) is strictly required by TPK-IIB, irrespective of peptide conformation, although a beta-turn may enhance the phosphorylation of those peptides that satisfy this requirement. Two other spleen tyrosine kinases, TPK-I/lyn and TPK-III, both related to the src family, also have a far greater preference for the peptide EDNEYTA over EPQYQPA. However, they can be distinguished from TPK-IIB by their lower affinity for the peptides EDNEYTA and NEYTA and by their different specificity towards the substituted derivatives of NEYTA. TPK-I/lyn, accepts most of the substitutions that are detrimental to TPK-IIB, the triply substituted peptide AAYAA being actually preferred over the parent peptide NEYTA. The substitution of glutamic acid by alanine is also tolerated by TPK-III, although, in contrast to TPK-IIB, the phosphorylation efficiency is drastically decreased by the substitution of the asparagine at position -2.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Peptides reproducing the phosphoacceptor sites of pp60c-src as substrates for TPK-IIB, a splenic tyrosine kinase devoid of autophosphorylation activity. 171 42

The peptide SAEEEDQYN, corresponding to the carboxyl-terminal tryptic fragment of rat progastrin, whose penultimate tyrosyl residue is sulphated in the native peptide, is phosphorylated with Km values of 120 and 180 microM by two spleen tyrosine protein kinases, termed TPK-IIB and TPK-III, respectively. Another spleen tyrosine protein kinase related to the src family (TPK-I/lyn) is poorly active toward this peptide, displaying a Km 6.5 mM. The Tyr-phosphorylated peptide is recognized by an antibody (L304), which reacts with both sulphated and unmodified peptides, while it is not recognized by a second antibody (L303), which reacts with unmodified peptide yet not with the sulphated derivative. These data, in conjunction with previous observations (Hofsteenge, J., Stone, S.R., Donella-Deana, A. and Pinna, L.A. (1990) Eur. J. Biochem. 188, 55-59) support the view that phosphotyrosine is an effective surrogate for sulphotyrosine in a wide spectrum of biological activities.
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PMID:Substitution of phosphotyrosine for sulphotyrosine in biologically active peptides. Enzymatic phosphorylation of a progastrin peptide confers immunoreactivity reminiscent of the sulphated derivative. 171 35

We have identified, purified, and characterized two neurofilament/tau kinases from bovine brain, PK36 and PK40, with apparent Mr of 36,000 and 40,000 and with novel biochemical properties. A specially designed immunoassay for phosphorylated epitopes in neurofilament (NF) proteins was used in the early stages of the purification. Neither kinase is closely associated with the cytoskeleton. Both kinases phosphorylate bovine intermediate (NF-M) and heavy (NF-H) NF subunits and also bovine tau at the expected KSP sequences, though other sites cannot be ruled out. In human paired helical filaments, tau, phosphorylated at these same KSP sites, is a major characterized constituent. Neither kinase is activated by the usual second messengers. Tau and the above NF subunits are phosphorylated in high stoichiometric ratios. In the intermediate NF subunit, all the expected sites appear to be phosphorylated, but in the heavy NF subunit only 7 out of the greater than 40 expected sites can be phosphorylated by our kinases. We demonstrate that both kinases can induce considerable shifts of apparent Mr with SDS-PAGE for tau and, for the first time in vitro, also for the intermediate NF subunit. Interestingly, PK36 and particularly PK40 are strongly inhibited by an excess of free ATP. We propose that during normal aging, and in Alzheimer's disease, age-related mitochondrial dysfunction would reduce ATP levels, which in turn might release the neurofilament/tau kinase from inhibition with consequent paired helical filament formation.
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PMID:Two novel kinases phosphorylate tau and the KSP site of heavy neurofilament subunits in high stoichiometric ratios. 171 59

In resting human epithelial and fibroblastic cells, c-Jun is phosphorylated on serine and threonine at five sites, three of which are phosphorylated in vitro by glycogen synthase kinase 3 (GSK-3). These three sites are nested within a single tryptic peptide located just upstream of the basic region of the c-Jun DNA-binding domain (residues 227-252). Activation of protein kinase C results in rapid, site-specific dephosphorylation of c-Jun at one or more of these three sites and is coincident with increased AP-1-binding activity. Phosphorylation of recombinant human c-Jun proteins in vitro by GSK-3 decreases their DNA-binding activity. Mutation of serine 243 to phenylalanine blocks phosphorylation of all three sites in vivo and increases the inherent trans-activation ability of c-Jun at least 10-fold. We propose that c-Jun is present in resting cells in a latent, phosphorylated form that can be activated by site-specific dephosphorylation in response to protein kinase C activation.
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PMID:Activation of protein kinase C decreases phosphorylation of c-Jun at sites that negatively regulate its DNA-binding activity. 184 81

A highly purified preparation of protein kinase FA (where FA is the activating factor for phosphatase 1)/glycogen synthase kinase 3 from rabbit muscle readily phosphorylated bovine neurofilaments. All three neurofilament proteins, the high, middle, and low molecular proteins (NF-H, NF-M, and NF-L), were phosphorylated when intact filaments were incubated with the kinase. Experiments with individual proteins showed that NF-M was the best substrate. At protein concentrations of 0.13 mg/ml, the initial rate of NF-M phosphorylation was 30% of that observed for glycogen synthase. Km values were 0.24 mg/ml (7 x 10(-7) M tetramer) for glycogen synthase and 0.10 mg/ml (5 x 10(-7) M dimer) for NF-M. Vmax values were 0.36 mumol/min/mg for glycogen synthase and 0.035 mumol/min/mg for NF-M. Dephosphorylated NF-M was phosphorylated only half as much as native NF-M; this is consistent with the known substrate specificity of the kinase. The possible involvement of FA/GSK-3 in the phosphorylation of neurofilaments in vivo is discussed.
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PMID:Phosphorylation of bovine neurofilament proteins by protein kinase FA (glycogen synthase kinase 3). 185 Jul 42

A novel brain-specific 25 kDa protein (p25) was purified from a bovine brain extract. The protein was phosphorylated by Ser/Thr-Pro kinase (TPK II) in tau protein kinase fractions at the Ser residues of Ser-Pro sequences. Using immunoblot analysis, the protein was found only in brain extracts, and was most abundant in the brain regions such as cerebrum and hippocampus, but less abundant in cerebellum, medulla oblongata and olfactory bulb. The protein was detected in rat, bovine and human brain extracts, indicating that this protein specifically exists in mammalian brain tissues.
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PMID:A novel brain-specific 25 kDa protein (p25) is phosphorylated by a Ser/Thr-Pro kinase (TPK II) from tau protein kinase fractions. 190 72

Hirudin, the powerful anticoagulant agent of leech (Hirudo medicinalis) saliva, was readily phosphorylated by two spleen tyrosine protein kinases (TPK-IIB and TPK-III) at Tyr63 with Km values of 238 microM and 74 microM, respectively. The synthetic tridecapeptide DGDFEEIPEEYLQ, corresponding to the hirudin 53-65 C-terminal fragment, was phosphorylated even more efficiently than hirudin itself. Four hirudin mutants, in which one or more of the glutamic acids at positions 57, 58, 61 and 62 have been replaced by glutamines, were poorer substrates than hirudin. The mutant in which all four glutamates were substituted, [Gln57,58,61,62]hirudin, was virtually not phosphorylated by either TPK-IIB and TPK-III. Substitution of Glu57 and Glu58 was less deleterious than substitution of the two glutamic acids adjacent to Tyr63: [Gln61,62]hirudin exhibited a 20-fold lower phosphorylation efficiency with TPK-IIB. With TPK-III, however, the Km value of [Gln61,62]hirudin was slightly lower, while the Vmax decreased sixfold. The substitution of Glu62 alone was also more detrimental with TPK-IIB than with TPK-III. The behaviour of a third spleen TPK, named lyn TPK-I and belonging to the src family, was markedly different in that it did not phosphorylate hirudin but exhibited significant activity towards [Gln57,58,61,62]hirudin. Taken together, these data confirm and extend with a protein substrate the results obtained with short model peptides which indicated the stringent substrate requirements of TPK-IIB (and of TPK-III to a lesser extent) for N-terminal acidic residues. In contrast, such residues are deleterious with lyn TPK-I. These observations also support the concept that tyrosine protein kinases recognize specificity determinants situated in the vicinity of the target residue rather than requiring higher-order structural features.
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PMID:Specificity determinants for tyrosine protein kinase. A study with recombinant hirudin mutants. 193 46

The previously isolated spleen tyrosine protein kinase, conventionally termed TPK-IIA, displaying activation by either positively or negatively charged polyelectrolytes has been further characterized. TPK-IIA is immunologically related with the tyrosine protein kinase encoded by the lyn gene, a member of src subfamily and is dramatically activated by very high NaCl concentration. The stimulatory effects of NaCl and polylysine, which are not additive, are accounted for by increased Vmax values, the Km being virtually unchanged, suggesting that both effectors probably interact with the same site(s). Stimulation of TPK-IIA by heparin appears to be partially additive to that promoted by NaCl and possibly occurring through a different mechanism. The NaCl activatory effect correlates with the electrolytic nature of synthetic peptides used as substrates, being much more consistent with neutral peptides as compared with acidic ones. Of the other three spleen tyrosine protein kinases, TPK-I shows similar biochemical and immunological features, suggestive of close relatedness with TPK-IIA, while TPK-IIB and TPK-III are neither related with the lyn protein nor with the products of three other oncogenes of the src subfamily, namely lck, hck and fyn.
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PMID:Stimulation by NaCl, polylysine and heparin of two forms of spleen tyrosine protein kinase immunologically related with the protein expressed by lyn oncogene. 199 63


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