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At least 919 records · Page 51Linked to original sources
PubMed1984

Alterations in hepatic protein kinase activity induced by triiodothyronine.

Quantitative and qualitative alterations in protein kinase (PK) activity in preparations of rat liver from hypothyroid (H) and T3-treated hypothyroid animals (T) were analyzed in comparison to enzyme from normal (N) animals. Qualitative variations in type of protein kinase were assessed by chromatography of cytosol preparations on DE-52 cellulose columns. Cytosol kinases resolved into a small fraction I containing a catalytic subunit, fraction II containing a type 1 holoenzyme, fraction III containing a cyclic-AMP independent PK, and fraction IV containing a type 2 holoenzyme. A cytosolic kinase active with casein as substrate was also identified. In H the type 1 holoenzyme was reduced in comparison to N. There were no other qualitative changes. Nuclear PKs were extracted with solvents containing 0.3 M KCl. Qualitative changes were evaluated by chromatography on phosphocellulose columns, but available methodology did not give reproducible evidence of changes in individual PKs. H had significantly more cytosolic PK active with protamine substrate than did normal animals, and by administration of T3, the H level was reduced progressively over 48 h to the N level. The activation ratio of total PK in cytosol was higher in H, and was also reduced to the N level by T3 administration. This suggests a higher steady state level of cyclic-AMP may be present in H rat liver cytosol. Cytosolic protein kinase reactive with casein as substrate increased gradually over 48 h after T3 administration from the H to the higher N level. It was significantly elevated to 110% of the H value by 48 h after T3 administration. The behavior of nuclear PK was entirely different. Nuclear PK reactive with protamine as substrate was increased within 1-1/2 h after T3 administration, reaching a peak of 130% of the control value at 5 h and returning to the normal level by 48 h. In contrast, nuclear PK reactive with casein as substrate also increased by 1-1/2 h after T3 administration and remained elevated at 110-115% of the control value throughout 48 h after T3 administration. The early changes in nuclear PK activity were prevented by administration of cycloheximide or alpha-amanitin. The observed changes in cytosolic PK, including the increment induced by T3 in type 1 enzyme, reduction in

Animals↗
PubMed1990

Infertility in male transgenic mice: disruption of sperm development by HSV-tk expression in postmeiotic germ cells.

Previous experiments revealed that male transgenic mice bearing a cosmid that included the Class II E alpha gene, about 35 kb of 5' flanking DNA, and the cosmid vector sequences were sterile. To ascertain the cause of the sterility, various subfragments of the cosmid were tested in transgenic mice. Only those pieces of DNA that included some of the E alpha flanking chromosomal DNA and the herpes simplex virus (HSV)-thymidine kinase (tk) gene that was in the vector resulted in male sterility. Histological analysis revealed abnormalities in nuclear morphology of elongating spermatids and retention of mature spermatids within the seminiferous epithelium. Immunocytochemical studies showed that the HSV-tk gene was expressed at low levels in postmeiotic round spermatids and at higher levels in more mature elongating spermatids. To determine whether expression of HSV-tk in spermatids might be responsible for the sterility, the protamine gene promoter was used to direct the expression of HSV-tk to postmeiotic germ cells. Since the mice so treated were also sterile, the data suggest that expression of this enzyme in spermatids is responsible for the sterility phenotype.

Animals↗
PubMed1990

Substrate-dependent inhibition of protein kinase C by specific inhibitors.

Protein kinase C (PKC) and its proteolysis-derived protein kinase independent of Ca2+ and phospholipids (PKM), were purified from rat brain. By using histone H1 and protamine as substrates, we assayed the effect of several inhibitors of PKC and PKM. The inhibition turned out to be dependent on both the nature of the kinase and the type of substrate assayed. These results may help to interpret the different responses elicited by PKC inhibitors in vivo.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1996

DNA-binding sperm proteins with oligo-arginine clusters function as potent activators for egg CK-II.

The stimulatory effect of DNA-binding sperm proteins (histone and protamine) on the phosphorylation of p98 (ERp99/GRp94, one of the Hsp-90 family of proteins) by egg casein kinase II (CK-II) was investigated in vitro. It was found that (i) phosphorylation of p98 by egg CK-II in vitro is greatly stimulated by poly-Arg, but not by poly-Lys; and (ii) similar stimulation is observed with sperm histones H2B2 and H2B3 (sea urchin) and fish protamines, such as salmine A1 (salmon) and protamine 3a (rainbow trout). These findings suggest that these DNA-binding sperm proteins function as potent activators for CK-II in fertilized eggs. All of these DNA-binding sperm proteins contain at least an oligo-Arg cluster as a common feature, which can interact with an acidic amino acid cluster of the regulatory beta-subunit CK-II.

Amino Acid Sequence↗
PubMed1986

Phosphorylation of the modulator protein of the ATP, Mg-dependent protein phosphatase by casein kinase TS. Reversal by PCS phosphatases and control by distinct phosphorylation site(s).

The phosphorylation by casein kinase TS (II) of the modulator protein of the ATP, Mg-dependent phosphatase increases after preincubation with the PCSH1 phosphatase or with the catalytic subunit of the ATP, Mg-dependent phosphatase. Dephosphorylation by the two phosphatases combined leads to the incorporation of 2 mol phosphate per mol modulator (at Ser residues). Occupancy of the ATP, Mg-dependent phosphatase phosphorylation site(s) is a negative determinant in the phosphorylation of the modulator by kinase TS. Among the PCS phosphatases PCSH1 shows the highest activity toward the 32P-Ser residues labeled by kinase TS in untreated or previously dephosphorylated modulator, while the ATP, Mg-dependent phosphatase is totally ineffective. Protamine stimulates all phosphatase activities, so that the catalytic subunit of the ATP, Mg-dependent phosphatase becomes almost as effective as the PCSC phosphatase in dephosphorylating the kinase TS sites.

Adenosine Triphosphate↗
PubMed1976

An adenosine 3':5' monophosphate dependent protein kinase from sea urchin spermatozoa.

A cyclic AMP dependent protein kinase (EC 2.7.1.37) from sea urchin sperm as purified to near homogeneity and characterized. A 68-fold purification of the enzyme was obtained. This preparation had a specific activity of 389 000 units/mg protein with protamine as the substrate. On the basis of the purification required, it may be calculated that the protein kinase constitutes as much as 1.5% of the soluble protein in sperm. There appeared to be a single form of the enzyme in sea urchin sperm, based on the behavior of the enzyme during DEAE-cellulose and Sephadex G-200 column chromatography. Magnesium ion was required for enzyme activity. The rate of phosphorylation of protamine was stimulated 2.5-fold by an optimal concentration of 0.9 M NaCl. The Km for ATP (minus cyclic AMP) was 0.119 +/- 0.013 (S.D.) and 0.055 mM +/- 0.009 (S.D.) in the presence of cyclic AMP. The specificity of the enzyme toward protein acceptors, in decreasing order of phosphorylation, was found to be histone f1 protamine, histone f2b, histone f3 and histone f2a; casein and phosvitin were not phosphorylated. The holoenzyme was found to have an apparent molecular weight of 230 000 by Sephadex G-200 chromatography. In the presence of 5 - 10(-6) M cyclic AMP, the holoenzyme was dissociated on Sephadex G-200 to a regulatory subunit of molecular weight 165 000 and a catalytic subunit of Mr 73 000. The dissociation could also be demonstrated by disc gel electrophoresis in the presence and absence of cyclic AMP.

Animals↗
PubMed1988

Protein kinase C activities with different characteristics, including substrate specificity, from two human HL-60 leukemia cell variants.

To evaluate the role of protein kinase C in the cellular maturation processes induced by phorbol diesters, we examined the biochemical activity of protein kinase C from HL-205, a cell variant from the human promyelocytic HL-60 leukemia that is susceptible to differentiation induced by phorbol 12-myristate 13-acetate, and from HL-525, an HL-60 variant that is resistant to such an induction. The activities of protein kinase C from the two cell types differed in their requirements for the cofactors Ca2+ and lipids. These enzyme activities also differed in their abilities to phosphorylate protamine and a series of four oligopeptides. We suggest that the differences in vitro in the activities of protein kinase C between HL-205 and HL-525 cells, especially in their substrate specificity, are closely related to the different phosphorylation patterns induced in vivo by phorbol 12-myristate 13-acetate in these cells. We also suggest that these differences may be responsible for the different susceptibilities of the two cell types to maturation induced by phorbol diesters.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1989

Autophosphorylation activates the soluble cytoplasmic domain of the insulin receptor in an intermolecular reaction.

The cytoplasmic protein-tyrosine kinase domain of the insulin receptor (residues 959-1355) has been expressed as a soluble protein in Sf9 insect cells via a Baculovirus expression vector (Ellis, L., Levitan, A., Cobb, M.H., and Ramos, P. (1988) J. Virol. 62, 1634-1639). The purified protein is a monomer as judged by its behavior in sucrose gradients and on gel filtration in the presence or absence of protamine. The initial rate of autophosphorylation using 3 mM MgCl2 is increased 20-30-fold by protamine. A maximum of 4-5 mol of phosphate are incorporated per mol of enzyme. The activity of the enzyme as a function of phosphorylation state was studied for three substrates: a synthetic dodecapeptide derived from the sequence of the major autophosphorylation site in the insulin receptor, poly(Glu, Tyr), 4:1, and histone 2B. Autophosphorylation of the protein to a stoichiometry of 4-5 mol of phosphate/mol increases its enzymatic activity as much as 200-fold; a 30-fold increase in activity occurs upon addition of 1 mol of phosphate/mol. The activities of unphosphorylated enzyme with the three substrates are 3.4, 2.3, and 0.44 nmol/min/mg, respectively. The activities of the autophosphorylated enzyme with the three substrates are 175, 274, and 45 nmol/min/mg, respectively. Exposure of the autophosphorylated enzyme to ADP results in a loss of phosphate from the enzyme which is associated with a decrease in enzymatic activity. Autophosphorylation of the kinase in the presence or absence of protamine displays a marked dependence on enzyme concentration. Furthermore, the rate of autophosphorylation decreases as the viscosity of the solution increases. Taken together, these data suggest that phosphorylation occurs via an intermolecular reaction.

Adenosine Diphosphate↗
PubMed1983

Purification and characterization of a phosvitin kinase from the thyroid gland.

1. Two cyclic AMP independent protein kinases phosphorylating preferentially acidic substrates have been identified in soluble extract from human, rat and pig thyroid glands. Both enzymes were retained on DEAE-cellulose. The first enzyme activity eluted between 60 and 100 mM phosphate (depending on the species), phosphorylated both casein and phosvitin and was retained on phosphocellulose; this enzyme likely corresponds to a casein kinase already described in many tissues. The second enzyme activity eluted from DEAE-cellulose at phosphate concentrations higher than 300 mM, phosphorylated only phosvitin and was not retained on phosphocellulose. These enzymes were neither stimulated by cyclic AMP, cyclic GMP and calcium, nor inhibited by the inhibitor of the cyclic AMP dependent protein kinases. 2. The second enzyme activity was purified from pig thyroid gland by the association of affinity chromatography on insolubilized phosvitin and DEAE-cellulose chromatography. Its specific activity was increased by 8400. 3. The purified enzyme (phosvitin kinase) was analyzed for biochemical and enzymatic properties. Phosvitin kinase phosphorylated phosvitin with an apparent Km of 100 micrograms/ml; casein, histone, protamine and bovine serum albumin were not phosphorylated. The enzyme utilized ATP as well as GTP as phosphate donor with an apparent Km of 25 and 28 microM, respectively. It had an absolute requirement for Mg2+ with a maximal activity at 4 mM and exhibited an optimal activity at pH 7.0. The molecular weight of the native enzyme was 110 000 as determined by Sephacryl S300 gel filtration. The analysis by SDS-polyacrylamide gel electrophoresis revealed a major band with a molecular weight of 35000 suggesting a polymeric structure of the enzyme.

Animals↗
PubMed1998

Purification and isotype analysis of protein kinase C from rat liver nuclei.

The properties and subtype composition of protein kinase C present in rat liver nuclei were studied in a Triton-X-100 extract of isolated purified nuclei. The enzyme activity was dependent on both Ca2+ and phosphatidylserine, but the phorbol ester 12-O-tetradecanoylphorbol 13-acetate gave only a partial stimulation. Both histone and myelin basic protein served as substrate. Purification of the Triton-X-100 extract followed by Q-Sepharose chromatography gave a preparation with a specific activity of 70 pmol/mg protein min. Western blotting of this preparation showed only the presence of the delta and zeta subtypes, but not the alpha-subtype, although the latter was present in rat liver homogenates. The beta, gamma and epsilon subtypes were not found in the homogenate nor in the nuclear extract. The specific activity of protein kinase C could be further increased up to 800 pmol/mg protein min after protamine agarose chromatography. Also in this preparation the presence of the delta and zeta subtypes could be established.

Amino Acid Sequence↗
PubMed1987

Evidence that the viral Ki-ras protein, but not the pp60v-src protein of ASV, stimulates proliferation through the PDGF receptor.

Protamine sulfate (PS), a specific blocker of PDGF action, inhibited the proliferative response of tsKSV-NRK cells to a reactivated, temperature-sensitive viral Ki-RAS protein, but it did not affect the proliferative response of tsASV-NRK cells to a reactivated pp60v-src protein kinase. The inhibition by PS of the proliferation response of tsKSV-NRK cells to reactivated Ki-RAS protein was overcome by serum growth factors, notably EGF, and concentrated serum-free conditioned medium from cultured NRK cells infected with wild-type KSV, but not by a combination of PDGF and insulin. These observations suggest that the viral Ki-RAS protein, but not pp60v-src, stimulates proliferation exclusively by inducing the host cells to produce PDGF or PDGF-like mitogenic factors.

Animals↗
PubMed1988

Biochemical characterization of rat brain protein kinase C isozymes.

Biochemical characteristics of three rat brain protein kinase C isozymes, types I, II, and III, were compared with respect to their protein kinase and phorbol ester-binding activities. All three isozymes appeared to be alike in their phorbol ester-binding activities as evidenced by their similar Kd for phorbol 12,13-dibutyrate and requirements for Ca2+ and phospholipids. However, differences with respect to the effector-mediated stimulation of protein kinase activity were detectable among these isozymes. The type I enzyme could be stimulated by cardiolipin to a greater extent than those of the type II and III enzymes. In the presence of cardiolipin, the concentrations of dioleoylglycerol or phorbol 12,13-dibutyrate required for half-maximal activation (A1/2) of the type I enzyme were nearly an order of magnitude lower than those for the type II and III enzymes. In the presence of phosphatidylserine, differences in the A1/2 of dioleoylglycerol and phorbol 12,13-dibutyrate for the three isozymes of protein kinase C were less significant than those measured in the presence of cardiolipin. Nevertheless, the A1/2 of these two activators for the type I enzyme were lower than those for the type II and III enzymes. At high levels of phosphatidylserine (greater than 15 mol %), binding of phorbol 12,13-dibutyrate to the type I enzyme evoked a corresponding stimulation of the kinase activity, whereas binding of this phorbol ester to the type II and III enzymes produced a lesser degree of kinase stimulation. For all three isozymes, the concentrations of phosphatidylserine required for half-maximum [3H]phorbol 12,13-dibutyrate binding were almost an order of magnitude less than those for kinase stimulation. Consequently, neither isozyme exhibited a significant kinase activity at lower levels of phosphatidylserine (less than 5 mol %) and phorbol 12,13-dibutyrate (50 nM), a condition sufficient to promote near maximal phorbol ester binding. In addition to their different responses to the various activators, the three protein kinase C isozymes also have different Km values for protein substrates. The type I enzyme appeared to have lower Km values for histone IIIS, myelin basic protein, poly(lysine, serine) (3:1) polymer, and protamine than those for the type II and III enzymes. These results documented that the three protein kinase C isozymes were distinguishable in their biochemical properties. In particular, the type I enzyme, which is a brain-specific isozyme, is distinct from the type II and III enzymes, both have a widespread distribution among different tissues.

Animals↗
PubMed2000

Mechanism of activation of cAMP-dependent protein kinase: in Mucor rouxii the apparent specific activity of the cAMP-activated holoenzyme is different than that of its free catalytic subunit.

Kinetic constants for peptide phosphorylation by the catalytic subunit of the dimorphic fungus Mucor rouxii protein kinase A were determined using 13 peptides derived from the peptide containing the basic consensus sequence RRASVA, plus kemptide, S6 peptide, and protamine. As a whole, although with a greater Km, the order of preference of the peptides by the M. rouxii catalytic subunit was similar to the one displayed by mammalian protein kinase A. Particularly significant is the replacement of serine by threonine in the basic peptide RRATVA, which impaired its role as a substrate of M. rouxii catalytic subunit. Mucor rouxii protein kinase A is a good model in which to study the mechanism of activation since cAMP alone is not enough to promote activation and dissociation. Four peptides were selected for the study of holoenzyme activation under conditions in which the enzymatic activity was not proportional to the holoenzyme concentration: RRASVA, RRRRASVA, KRRRLSSRA (S6 peptide), and LRRASLG (kemptide); protamine was used as reference. Differential activation degree was observed depending on the peptide used and on cAMP concentration. Ratios of activity between different substrates displayed by the holoenzyme under the above conditions did not reflect the one expected for the free catalytic subunit. The degree of inhibition of the holoenzyme activity by an active peptide derived from the thermostable protein kinase inhibitor was dependent on the substrate used and on the holoenzyme concentration, while it was found to be independent of these two parameters for free catalytic subunit. Polycation modulation of holoenzyme activation by cAMP was also dependent on the polycation itself and on the peptide used as substrate. The observed kinetic differences between holoenzyme and free catalytic subunit were decreased or almost abolished when working at low enzyme or at high cAMP concentrations. Two hypotheses compatible with the results are discussed: substrate participation in the dissociation process and/or holoenzyme activation without dissociation.

Amino Acid Sequence↗
PubMed1975

Regulation of C3-enzymes in facultative phototrophic bacteria: the cold-labile pyruvate kinase of Rhodopseudomonas sphaeroides.

Pyruvate kinase (EC2.7.1.40) from Rhodopseudomonas sphaeroides was purified 40-fold be precipitation with protamine sulfate and ammonium sulfate followed by gel-filtration. The preparations obtained from cells grown with different carbon sources or cultural conditions differ with respect to specific activity but not with respect to molecular weight (250000 dalton) or regulatory properties. The phosphoenolpyruvate (PEP)-saturation cruve of the enzyme is sigmoidal with Hill coefficients varying from nH equals 1.8 (pH 9.2) to 2.7 (pH 6.0). The enzyme is activated by adenosinemonophosphate (AMP) and the sugarmonophosphates ribose-5-phosphate (R-5-P), glucose-6-phosphate (G-6-P), and-to a lesser extent-fructose-6-phosphate (F-6-P). Fructose-1.6-bisphosphate (FDP) has no measurable effect. Inhibitors of the enzyme are adenosinetriphosphate (ATP), inorganic phosphate (Pi) and the dicarboxylic acids succinate and fumarate. Kinetic analysis reveals that the sugar-phosphates and the dicarboxylic acids act as true allosteric ligands, whereas the effects of AMP, ATP, and Pi cannot be interpreted soley in terms of allosteric interactions. Cold-treatment of the enzymes lead to a rapid loss of activity, but does not change the regulatory properties of the enzyme. Analysis of the kinetics of cold-inactivation and its reversal at 30 percent C, together with studies on the gelfiltration behaviour of the native and the cold-treated enzyme make it likely that the cold-induced loss of activity is due to a dissociation of the enzyme.

Adenosine Monophosphate↗
PubMed1985

Manganese-phospholipid-stimulated protein kinase activity of human leukemic cells.

A protein kinase system with unusual characteristics was noted in extracts of HL-60 cells using endogenous proteins as substrates. This system exhibited a cation preference for manganese at an optimal concentration of 0.5 mM. Moderate activity was detectable with magnesium at an optimal concentration of 5.0 mM, but calcium was inactive. Activity was markedly stimulated by phospholipid, with phosphatidylglycerol and phosphatidylinositol exhibiting greater activity than phosphatidylserine. In isolated plasma membranes, the major substrate of this system was a 73-kDa protein, while cytoplasmic extracts exhibited larger amounts of a 42-kDa substrate. 73-kDa phosphorylating activity of membranes was comparably active at 0 and 31 degrees C, although in cytosol activity was greater at 31 degrees C. No 73-kDa protein phosphorylation was observed in the presence of Ca2+, Mg2+, and phosphatidylserine. Phosphoamino acid analysis of the 73-kD band revealed phosphothreonine and phosphoserine. The 42-kDa substrate was distinguishable from actin by two-dimensional gel electrophoresis, which disclosed that both major substrates were highly basic in the isoelectric focusing dimension. Protamine and histones (H2B greater than H1 greater than H3) exhibited phospholipid-stimulated phosphorylation in the presence of Mn2+, but phosvitin, casein, and vinculin were not substrates. High levels of phosphorylative activity involving the 73-kDa substrate were noted in nuclear extracts. Complex patterns of increase of this activity were noted in both cytosol and nuclear extracts following induction of differentiation with dimethyl sulfoxide, retinoic acid, or phorbol 12-myristate 13-acetate. This study thus demonstrated the presence of a previously undescribed type of protein kinase activity which exhibited alterations during leukemic cellular differentiation.

Cell Differentiation↗
PubMed1997

Activation of protein kinase C subtypes alpha, gamma, delta, epsilon, zeta, and eta by tumor-promoting and nontumor-promoting agents.

Protein kinase C (PKC) subtypes alpha, gamma, delta, epsilon, zeta, and eta have been expressed using the baculovirus expression system. The partially purified PKC subtypes have been studied for their substrate specificities and phospholipid-independent activation by various chemically different nontumor- and tumor-promoting agents, as well as their inhibition of kinase activity by staurosporine and two related compounds. An endogenous PKC-like kinase activity of Sf9 cells was detected and analyzed for cofactor requirements and inhibition. Protamine sulfate was most efficiently phosphorylated by all of the PKC subtypes tested, although this phosphorylation was independent of phosphatidylserine (PS) and diacylglycerol (DAG) or 12-O-tetradecanoylphorbol 13-acetate (TPA). Except for PKC-zeta, all subtypes tested phosphorylated myelin basic protein (MBP), histone, or a peptide derived from the pseudosubstrate region of PKC-alpha in a PS/DAG-dependent manner but to varying extents. Among the various agents tested, TPA most efficiently stimulated the kinase activities of the PKC subtypes in a phospholipid-dependent manner. Phorbol 12,13-dibutyrate (PDBu) was less effective than TPA but displayed no major difference among the subtypes. Activation of PKC-alpha by bryostatin-1 reached only half of the TPA response whereas the other subtypes were activated more effectively. The weak tumor promoter resiniferonol 9,13,14-orthophenyl acetate (ROPA) mainly stimulated PKC-alpha and PKC-gamma at 1 microM concentration, whereas PKC-epsilon and PKC-eta were much less activated. Sapintoxin D, mezerein, indolactam V, and resiniferatoxin at concentrations of 1-100 nM preferentially activated PKC-alpha in a DAG-like manner, whereas at 1 microM other subtypes were activated as well. Preferential activation of PKC-alpha was also noted for tinyatoxin and thapsigargin, but their mode of activation is unclear because these two compounds did not compete for the phorbol ester binding of the PKC subtypes as the other agents did. Of the three PKC inhibitors tested, staurosporine most efficiently inhibited kinase activity of the PKC subtypes, whereas K252a and CGP 41251 were at least 10 times less effective. However, K252a showed certain specificity for inhibition of PKC-alpha, and CGP 41251 failed to inhibit PKC-epsilon and PKC-zeta. Given the different substrate specificities and modes of activation by various tumor-promoting and nontumor-promoting agents, as well as the different sensitivities towards different inhibitors, our results indicate a divergence of individual PKC subtypes in signal transduction.

Animals↗
PubMed1983

[Characterization of cyclic AMP-dependent protein kinase from smooth muscle of cattle and its modification by drugs].

Cyclic AMP-dependent protein kinase activity (ATP: protein phosphohydrolase, EC 2.7.1.37) from the postmicrosomal supernatant (100 000 X g supernatant) of bovine coronary and carotid arteries was isolated by ammonium sulfate precipitation followed by ion-exchange chromatography on DEAE-cellulose and partially characterized. Kinase activity was assayed in a standard system with [gamma-32P]-ATP and protamine at pH 7.5, 310 K, in the presence and absence of cyclic AMP. The apparent Km-values for ATP, Mg2+ and cAMP are 6.1 X 10(-5) M, 6.4 X 10(-4) M, 3.4 X 10(-8) M, respectively (coronary arteries) or 5.0 X 10(-5) M, 1.02 X 10(-3) M, 1.7 X 10(-8) M, respectively (carotids). Optimal concentrations of cAMP produced a 2-3-fold increase in the cyclic AMP-dependent protein kinase activity. After purification on DEAE-cellulose three peaks of cyclic AMP-dependent protein kinase activity were found. Especially the second peak fraction showed high protein kinase activity, strong [3H]cAMP-binding and high sensitivity to cAMP. Several drugs tested on the crude enzyme preparation showed stimulating or inhibitory activities in the concentration range of 1-10 microM.

Adenosine Triphosphate↗