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)

Cyclic GMP-dependent protein kinase was purified from foetal calf hearts, and its general properties and subunit structure were studied. The enzyme was purified over 900-fold from the heart extract by pH 5.3-isoelectric precipitation, DEAE-cellulose chromatography, Sephadex G-200 filtration and hydroxyapatite treatment. The purified myocardial enzyme, free from cyclic AMP-dependent protein kinase contamination, exhibited an absolute requirement of stimulatory modulator (or crude modulator containing the stimulatory modulator component) for its cyclic GMP-stimulated activity. Inhibitory modulator (protein inhibitor) of cyclic AMP-dependent protein kinase could not stimulate nor inhibit the cyclic GMP target enzyme. The enzyme had Ka values of 0.013, 0.033 and 3.0 micronM for 8-bromo cyclic GMP, cyclic GMP and cyclic AMP respectively. The cyclic GMP-dependent enzyme required Mg2+ and Co2+ for its activity, with optimal concentrations of about 30 and 0.5 mM respectively. The pH optimum for the enzyme activity ranged from 6 to 9. Histones were generally effective substrate proteins. The enzyme exhibited a greater affinity for histones than did the cyclic AMP-dependent class of protein kinase. The holoenzyme (apparent mol.wt. 150 000) of the myocardial cyclic GMP-dependent protein kinase was dissociated into a cyclic GMP-independent catalytic subunit (apparent mol.wt. 60 000) by cyclic GMP and histone. The catalytic subunit required the stimulatory modulator for its activity, as in the case of the holoenzyme in the presence of cyclic GMP.
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PMID:Guanosine cyclic monophosphate-dependent protein kinase from foetal calf heart. Purification, general properties and catalytic subunit. 1 43

A regulatory role for adenosine 3',5'-monophosphate (cyclic AMP) in the production of the renal hormone rythropoietin following erythropoietic stimulation with cobaltous chloride hexahydrate is proposed. Studies in rates reveal a temporal relationship between renal cyclic AMP levels and plasma titers of erythropoietin. In addition, cobalt increases the activity of an erythropoietin-generating enzyme (renal erythropoietic factor) with maximal enzyme activity occurring after the rise in cyclic AMP levels but before the increase in erythropoietin titers. This increase in renal cyclic AMP is localized to the renal cortex. Cobalt stimulates renal cortical adenylate cyclase but has no effect on renal cyclic nucleotide phosphodiesterase. The addition of cyclic AMP (3 time 10-6 M) and a partially purified cyclic AMP-dependent protein kinase from rat kidney to an inactive preparation of renal erythropoietic factor increases the ability of renal erythropoietic factor to generate erythropoietin. Data from the polycythemic mouse assay, a bioassay used to quantitate erythropoietic activity of test substances, indicate that dibutyryl cyclic AMP is erythropoietically active with respect to its ability to increase radioactive-labelled iron (59Fe) incorporation into heme of newly formed red blood cells. Theophylline, which by itself is erythropoietically inactive, potentiated the erythropoietic effect of cobalt in polycythemic mice. These results suggest that cyclic AMP plays a significant role in the renal production of erythropoietin following cobalt administration. It is postulated that cobalt stimulates renal cortical adenyoate cyclase, thus increasing renal cyclic AMP levels. Cyclic AMP then activates a protein kinase which subsequently stimulates renal erythropoietic factor to generate erythropoietin. A similar cyclic AMP mechanism may be operative after erythropoietic stimulation by exposure to hypoxia or prostaglandin treatment.
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PMID:The role of renal adenosine 3',5'-monophosphate in the control of erythropoietin production. 16 77

Guanosine 3':5'-monophosphate (cyclic GMP)-dependent protein kinase was purified from the guinea pig fetal lung, a tissue shown to be the richest in this enzyme in all mammalian sources examined, and its general properties studied. The enzyme was purified 150-fold from crude extract by steps of pH 5.4 isoelectric precipitation, Sephadex G-200 filtration, hydroxylapatite treatment and DEAE-cellulose chromatography. The purified enzyme, free from contamination with adenosine 3':5'-monophosphate (cyclic AMP)-dependent protein kinase, had a specific activity at least equivalent to 600-fold purification of the enzyme from the adult lung. The pulmonary enzyme exhibited an absolute requirement of protein kinase modulator (prepared from various mammalian tissues with an exception of skeletal muscle) for its activity. Inhibitor protein of cyclic AMP-dependent protein kinase purified from rabbit skeletal muscle could not stimulate nor inhibit the cyclic GMP target enzyme, indicating the factors from mammalian sources regulating the two classes of protein kinases may not be the same. The enzyme had Ka values of 1.3 times 10(-8) and 3.3 times 10(-8) M for 8-bromo cyclic GMP and cyclic GMP, respectively, compared to 3.0 times 10(-6) M for cyclic AMP. Cyclic GMP lowered the Km of the enzyme for ATP from 6.3 times 10(-5) M in its absence to 2.1 times 10(-5) M in its presence, accompanied by an approximate doubling of the Vmax. The molecular weight of the enzyme (assayed by its catalytic and cyclic GMP-binding abilities) was estimated to be 123,000, corresponding to a sedimendation coefficient of 7.06 S, by means of sucrose density gradient ultracentrifugation. The cyclic GMP-dependent enzyme required Mg2+ and Co2+ for its activity with optimal concentrations of about 30 and 0.7 mM, respectively. The maximal activity seen in the presence of Mg2+, however, was nearly twice as high as that seen in the presence of Co2+. Histones were generally effective substrates for the enzyme, whereas protamine, casein, phosvitin, phosphorylase kinase, and activator protein of phosphodiesterase were not. The cyclic GMP-dependent enzyme exhibited a greater affinity for histones than did the cyclic AMP-dependent enzyme in the presence of Mg2+.
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PMID:Purification and general properties of guanosine 3':5'-monophosphate-dependent protein kinase from guinea pig fetal lung. 17 61

A nucleoside-dependent protein kinase (EC 2.7.1.37) was partially purified from Trypanosoma gambiense, the pathogenic agent of sleeping sickness. This enzyme catalyzes the phosphorylation of histone and protamine. Various nucleosides at the concentration of 10(-4) M stimulated the histone kinase activity about two-fold, whereas cyclic AMP and cyclic GMP were without effect. The pH-optimum for histone phosphorylation was at about pH 7.0. The enzyme activity absolutely depends on Mg2+, Mn2+ or Co2+. The apparent Km-value for histone was 0.3 mg/ml and those for ATP were 2 - 10(-4) M and 6 - 10(-5) M in the absence or presence of 10(-4) M adenosine respectively. IDP and ADP complete with ATP. The inhibition constants were calculated to be 2 - 10(-4) M and 2.5 - 10(-4) M, respectively. The molecular weight of the histone kinase was found to be 95 000 by gel filtration and 88 000 by sedimentation in a sucrose gradient.
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PMID:Nucleoside-dependent protein kinase from Trypanosoma gambiense. 17 63

cGMP-dependent protein kinase from bovine lung has been purified to homogeneity using 8-(2-aminoethyl)-amino adenosine 3':5'-monophosphate/Sepharose. Conditions for adsorption of holoenzyme to the affinity chromatography media followed by competitive ligand elution with cGMP have been determined. The holoenzyme of 150,000 molecular weight is composed of two 74,000 molecular weight subunits which are linked in part by disulfide bridges. Two moles of cGMP are bound per mol of holoenzyme compatible with 1 mol of cGMP/monomer. Dissociation of subunits does not occur upon cGMP binding and protein kinase activation. cGMP-dependent protein kinase has an isoelectric point of 5.4 and a Stokes radius of 50 A. The enzyme is asymmetric with an f/f0 of 1.42 and an axial ratio of 7.4. Determination of enzyme activity at varying concentrations of ATP revealed that cGMP increased the Vmax for ATP without significant effect on the Km. The purified enzyme was maximally active at 5 mM Mg2+; other divalent cations could not substitute for Mg2+. In the presence of Mg2+, strong inhibitory effects of other cations were observed with Mn2+, greater than Zn2+, greater than Co2+ greater than Ca2+. Although maximal cGMP-dependence was observed at pH 5.7 to 7.0, basal activity rose at higher pH values to approach activity observed with cGMP. A molecular model comparing cGMP-dependent protein kinase with cAMP-dependnet protein kinase is presented.
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PMID:Guanosine 3':5'-monophosphate-dependent protein kinase from bovine lung. Subunit structure and characterization of the purified enzyme. 19 91

Plasma membrane fractions I and II isolated from bovine corpus luteum contain phosphoprotein phosphatases. Enzyme activities associated with both membrane fractions showed pH optima in the neutral range and were most active with phosphoprotamine as the exogenous substrate. The enzyme activity was partially inhibited by Co2+, Zn2+ and Fe2+. Dithioerythritol, glutathione (reduced) and 2-mercaptoethanol stimulated the enzyme activity, whereas N-ethylmaleimide and N-phenylmaleimide were inhibitory. Similarly, various cyclic nucleotides and nuclsoside triphosphates also inhibited phosphoprotein phosphatase activities. The phosphatase activity was also observed with endogenous phosphorylated membrane proteins as substrate. The endogenous phosphorylation of membranes was rapid and attained a maximal level after 15--20 min of incubation. Initially endogenous dephosphorylation was also very rapid, but did not reach completion. In addition to phosphoprotein phosphatase, membrane preparations also possessed very active cyclic-AMP-dependent protein kinase activity. Phosphoprotein phosphatase activity from plasma membranes was solubilized by ionic and nonionic detergents. Optimal solubilization was achieved with 0.1% sodium deoxycholate. Sucrose density gradient centrifugation of deoxycholate-solubilized fraction I and fraction II membranes resolved phosphoprotein phosphatase activity into two species with apparent sedimentation coefficients of 6.7 S (Mr 130000) and 4.8 S (Mr 90000). Cyclic-AMPstimulated protein kinase activity sedimented as a broad peak with a sedimentation coefficient of 5.5 S (Mr 110000).
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PMID:Solubilization and characterization of phosphoprotein phosphatase(s) from bovine corpus-luteum plasma membranes. 24 Jun 98

TRH and lysine-bradykinin (Lys-bradykinin) increase PRL release and arachidonate liberation from anterior pituitary cells. We investigated whether the arachidonate liberation stimulated by TRH and Lys-bradykinin originates in pituitary lactotropes and whether these events are accomplished through similar mechanisms. Lys-bradykinin and TRH rapidly (0.5 min) increased the intracellular [3H]arachidonate content of rat anterior pituitary cells. Lys-bradykinin also increased [3H]arachidonate liberation and PRL release from lactotrope-enriched pituitary cells, but not from a pituitary cell preparation with a diminished number of lactotropes. In contrast, TRH increased [3H]arachidonate liberation from both lactotrope-enriched and lactotrope-diminished preparations; this increased [3H]arachidonate liberation stimulated by TRH in the lactotrope-diminished cells may originate in the thyrotropes. The effects of TRH and Lys-bradykinin on [3H]arachidonate and [14C]stearate liberation in perfused pituitary cells also were determined. Both secretagogues increased arachidonate and stearate liberation in a biphasic manner, characterized by a transient spike, followed by a lower magnitude wave of fatty acid release. The spike phase produced by Lys-bradykinin was more pronounced than that produced by TRH. The calcium dependence of TRH- and Lys-bradykinin-stimulated arachidonate liberation also was investigated. Cobalt and the low calcium medium containing ionomycin were used to block the secretagogue-induced increase in intracellular calcium concentrations. These conditions blocked TRH-stimulated arachidonate liberation, but only marginally decreased Lys-bradykinin-stimulated arachidonate liberation, indicating that the two peptides act through different mechanisms. Therefore, TRH stimulation of arachidonate liberation is linked to an increase in intracellular calcium. In contrast, Lys-bradykinin increases arachidonate liberation through a calcium-independent intracellular mediator. This calcium-independent increase in arachidonate liberation may involve the bradykinin receptor being coupled directly to a phospholipase, a G-protein that provides a link between the bradykinin receptor and the phospholipases that liberate arachidonate, or bradykinin-induced activation of a protein kinase-C that activates the phospholipases and subsequently liberates arachidonate.
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PMID:Thyrotropin-releasing hormone and lysine-bradykinin stimulate arachidonate liberation from rat anterior pituitary cells through different mechanisms. 150 63

The functional significance of the oxidation/reduction state of sulfhydryl groups of cGMP-dependent protein kinase (cGMP kinase) was studied at 30 degrees C using different metal ions as oxidizing agents. Mn2+, Zn2+, Fe2+, Ni2+, and Co2+ failed to activate cGMP kinase, whereas Cu2+, Cu+, Fe3+, Hg2+, and Ag+ activated cGMP kinase by oxidation with an activity ratio (-cGMP/+cGMP) of about 0.7. The activation was not caused by degradation of the enzyme to a cGMP-independent constitutively active form. Reduction of the Cu(2+)-activated and gel-filtered enzyme with dithiothreitol lowered the activity ratio in the absence of cGMP to 0.17. Oxidation did not change the kinetic and binding parameters of cGMP kinase significantly but reduced the number of titratable sulfhydryl groups from 9.5 +/- 0.7 to 6.0 +/- 0.4 cysteines/75-kDa subunit. The free cysteinyl residues of the native and Cu(2+)-oxidized cGMP kinase were labeled with 4-dimethylaminoazobenzene-4'-iodoacetamide or N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide. Tryptic peptides of the labeled proteins were isolated and sequenced. The cysteinyl residues oxidized by Cu2+ were identified as disulfide bonds between Cys-117 and Cys-195 and Cys-312 and Cys-518, respectively. Cu2+ activation of cGMP kinase was prevented by mild carboxymethylation of the reduced enzyme with iodoacetamide, which apparently modified these four cysteinyl groups. The results show that cGMP kinase is activated by the formation of at least one intrachain disulfide bridge.
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PMID:Oxidation of cysteines activates cGMP-dependent protein kinase. 165 29

alpha-D-Mannosidase that exhibits a pH optimum close to neutrality (neutral mannosidase) purified from monkey brain cytosol is known to be stimulated by Co2+ and this stimulation is suggested to be mediated through a Co2+ activated aminopeptidase that is inseparable from the neutral mannosidase and that cleaves amino acids from the neutral mannosidase. In the present studies, the phosphorylation on serine residues of the neutral mannosidase by cyclic AMP dependent protein kinase is demonstrated. After phosphorylation the mannosidase activity remained unchanged, but it was not stimulated by Co2+. The aminopeptidase activity, although it retained its response to stimulation by Co2+, showed a drastic reduction in its activity after phosphorylation. It is suggested that the loss of Co2+ sensitivity of the neutral mannosidase after phosphorylation is mediated through the aminopeptidase.
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PMID:Loss of Co2+ sensitivity of monkey brain cytosolic neutral alpha-D-mannosidase by cyclic AMP dependent phosphorylation. 166 19

1. Bull-frog sympathetic neurones in primary culture were voltage clamped in the whole-cell configuration. The pipette solution contained ATP (5 mM). 2. A hyperpolarization-activated sodium-potassium current (H-current: IH) was separated from other membrane currents in a nominally calcium-free solution containing cobalt (2 mM), magnesium (4 mM), barium (2 mM), tetraethylammonium (20 mM), tetrodotoxin (3 microM), apamin (30 nM) and 4-aminopyridine (1 mM). IH was selectively blocked by caesium (10-300 microM). 3. The steady-state activation of IH occurred between -60 and -130 mV. The H-conductance was 4.1-6.6 nS at the half-activation voltage of -90 mV. With the concentrations of potassium and sodium ions in the superfusate at 20 and 70 mM, respectively, the reversal potential of IH was about -20 mV. IH was activated with a time constant of 2.8 s at -90 mV and 22 degrees C. The Q10 between 16 and 26 degrees C was 4.3. 4. A non-hydrolysable ATP analogue in the pipette solution did not support IH activation. Intracellular 'loading' of GTP-gamma-S (30-500 microM) led to a progressive activation of IH. 5. Forskolin (10 microM) increased the maximum conductance of IH by 70%. This was associated with a depolarizing shift in the half-activation voltage (5-10 mV) and in the voltage dependence of the activation/deactivation time constant of IH. 6. Essentially the same results as with forskolin were obtained by intracellular 'loading' with cyclic AMP (3-10 microM) or bath application of 8-bromo cyclic AMP (0.1-1 mM), dibutyryl cyclic AMP (1 mM) and 3-isobutyl-1-methylxanthine (0.1-1 mM). 7. The protein kinase inhibitor H-8 (1-10 microM) decreased the peak amplitude of IH. Phorbol 12-myristate 13-acetate (10 microM), a protein kinase C activator, was without effect. 8. It is concluded that a voltage-dependent cation current can be regulated by the basal activity of adenylate cyclase, presumably through protein kinase A, in vertebrate sympathetic neurones.
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PMID:Cyclic AMP regulates an inward rectifying sodium-potassium current in dissociated bull-frog sympathetic neurones. 169 Dec 92


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