Inhibition of foetal pulmonary choline-phosphate cytidylyltransferase under conditions favouring protein phosphorylation

Choline-phosphate cytidylyltransferase (EC 2.7.7.15) activity from 25- and 29-day-foetal rabbit lungs was inhibited in both the cytosolic and the microsomal fractions by preincubation with MgATP. The inhibition of the cytosolic enzyme was greater when measured with added phosphatidylglycerol (PG) than without (78-89% versus 50-55%), whereas the inhibition of the microsomal enzyme did not exhibit this distinction (66-72% versus 60-70%). When preincubated with the buffer alone, the cytosolic enzyme was activated to a greater extent by added PG than was the microsomal enzyme (13-14-fold versus 2-3-fold). However, after preincubation with MgATP, the cytosolic enzyme was activated to a smaller extent by added PG (3-6-fold). The inhibition of the enzyme by MgATP required a preincubation and was absent when ADP or AMP was substituted for ATP. Moreover, ATP analogues such as adenosine 5′-[beta, gamma-methylene]triphosphate and adenosine 5′-[γ-thio]triphosphate also failed to inhibit the enzyme when substituted for ATP in the preincubation. The inhibition by MgATP was not affected by including cyclic AMP in the preincubation, but Ca2+ ions alone or plus diacylglycerol in the preincubation increased the inhibition slightly. The inhibition was abolished by including an inhibitor of cyclic-AMP-dependent protein kinase in the preincubation. These observations, taken collectively, point to the inhibition of foetal pulmonary cytidylyltransferase through the phosphorylation of a protein and suggest that this key enzyme in lung surfactant production may be regulated through this mechanism.

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The amounts of rat liver cyclic AMP-dependent protein kinase I and II are differentially regulated by diet

Biochemical Journal ◽

10.1042/bj2560447 ◽

1988 ◽

Vol 256 (2) ◽

pp. 447-452 ◽

Cited By ~ 6

Author(s):

R Ekanger ◽

O K Vintermyr ◽

S O Døskeland

Keyword(s):

Protein Kinase ◽

Cyclic Amp ◽

Protein Restriction ◽

Type I ◽

Short Term ◽

Rat Hepatocyte ◽

Dependent Protein Kinase ◽

Key Enzyme ◽

Dependent Protein ◽

Fasted Rats

1. The fluctuations in rat hepatocyte volume and protein content in response to dietary perturbations (starvation, protein restriction, refeeding) were accompanied by corresponding fluctuations in the amount of the regulatory (R) and catalytic (C) subunits of cyclic AMP-dependent protein kinase. Thus the intracellular concentration of this key enzyme was adjusted to be near constant. 2. The adjustment of cellular R was accomplished almost exclusively by regulating cytosolic RI (R subunit of type I kinase). The preferential down-regulation of cytosolic RI in response to starvation/protein restriction indicates that particulate RI and cytosolic as well as particulate RII are more resistant to breakdown during general catabolism in the hepatocyte. 3. The diet-induced fluctuations of kinase subunits were uniformly distributed in all populations of parenchymatous hepatocytes, regardless of their size and density. It is thus possible to isolate hepatocytes with uniformly altered RI/RII ratio from livers of rats with different feeding regimens. 4. The binding of endogenous cyclic AMP to RI and RII was similar in livers with high RI/RII ratio (fed rats) and low RI/RII ratio (fasted rats) as well as in hepatocytes isolated from fasted rats. Under the conditions of the experiment (short-term stimulation by glucagon), therefore, neither the dietary state nor the RI/RII ratio seemed to affect the apparent affinity of the isoreceptors for cyclic AMP. However, RI appeared to show a slightly higher co-operativity of intracellular cyclic AMP binding than did RII in all states.

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Adenosine-3′,5′-Monophosphate-Dependent Protein Kinase From Bovine Anterior Pituitary Gland. III. Structural Specificity of the ATP Site of the Catalytic Subunit

Canadian Journal of Biochemistry ◽

10.1139/o74-021 ◽

1974 ◽

Vol 52 (2) ◽

pp. 137-141 ◽

Cited By ~ 19

Author(s):

Simon Lemaire ◽

Fernand Labrie ◽

Marie Gauthier

Keyword(s):

Protein Kinase ◽

Cyclic Amp ◽

Catalytic Subunit ◽

Protein Kinase Activity ◽

Dependent Protein Kinase ◽

Stringent Requirement ◽

Dependent Protein ◽

Atp Analogues ◽

High Concentrations ◽

Derivatives Of

The effect of analogues and derivatives of adenosine 3′,5′-monophosphate (cyclic AMP) and of ATP on the incorporation of 32P from [γ-32P]ATP into histones has been measured using the purified catalytic subunit of adenohypophyseal protein kinase. Gamma-labeled CTP, GTP, and UTP cannot substitute for [γ-32P]ATP but they slightly inhibit the phosphorylation by [γ-32P]-ATP when present as unlabeled compounds. A stringent requirement of the adenine nucleus is observed for the ability to compete at the ATP site, inhibitions of 42, 39, 32, and 63% being observed respectively with adenine, adenosine, 5′AMP, and ADP, while the corresponding purine or pyrimidine derivatives have no effect when present at a 13-fold molar excess relative to [γ-32P]ATP. The N6-benzoyl and N6-butyryl derivatives of cyclic AMP are inactive whereas the 8-substituted derivatives are generally as active as cyclic AMP itself, except for the 8-amino- and 8-hydroxy-derivatives, which exhibit a lower degree of competition. All cyclic AMP and ATP analogues and derivatives that inhibit histone phosphorylation by [γ-32P]ATP act as competitive inhibitors. Such competition at the ATP site of the catalytic subunit of protein kinase probably accounts for the progressive inhibition of cyclic-AMP-dependent protein kinase activity measured at high concentrations (above 10−5 M) of the cyclic nucleotide.

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