Phosphorylase kinase activity

During muscle contraction, l Caz+ is released from the sarcoplasmic reticulum. The Caz+ binds to the calmodulin subunit of phosphorylase kinase, activating it without phosphorylation. Phosphorylase kinase can then activate glycogen phosphorylase, causing glycogen degradation. 

Figure 18.14 Glycogen biosynthesis and degradation regulation. cAMP activates cAMP-dependent protein kinases. They cause the phosphorylation of glycogen synthase (inactivation), phosphorylase kinase (activation), and the inhibitory protein. The last inhibits phosphoprotein phosphatase. Activated phosphorylase kinase causes the phosphorylation of phosphorylase b, thus activating it to phosphorylase a. Phosphoprotein phosphatase is inhibited by the phosphorylated inhibitor protein. Such inhibition is released when the inhibitor protein is dephosphorylated. The phosphatase then reactivates glycogen synthase and inactivates phosphorylase kinase and phosphorylase a.

Ca + is required for phosphorylase activation in fat bodies of both P, americana (25) and discoidalis (personal observation). Addition of Ca + elevates fat body phosphorylase kinase activity in P. americana (33). and calmodulin inhibitors suppress CC-stimulated trehalose production by the fat body in vitro. However, direct addition of calmodulin to fat body phosphorylase kinase also suppresses the kinase activity. It is proposed that Ca + interacts directly with a calmodulin-like subunit of phosphorylase kinase to activate the enzyme, and the presence of exogenous calmodulin competes with the enzymic subunit for available Ca + (33). These results suggest that the HGHs may influence adipocyte Ca + levels related to phosphorylase activation to promote glycogenolysis for trehalose synthesis. Possibly, HGH-mediated fat body Ca levels may interact with polyphosphoinositides, diacyl glycerol and protein kinase C as second messengers for endocrine message transduction and phosphorylase activation. 

The rate of activation of phosphorylase is measured at pH 6.0 or 6.8 and pH 8.2, and the result is expressed as the ratio of phosphorylase kinase activity measured at the low pH to that measured at the high pH. An increase in the activity ratio of phosphorylase kinase indicates transformation to the activated (phosphorylated) form of the enzyme. Activated phosphorylase kinase is 25 to 50 times more active than inactive phos-... 

Phosphorylase kinase activity in dog heart homogenate has been measured by incubation at pH 6.0 and 8.2 with ATP, Mg, and crystalline phosphorylase b [135]. The h to a conversion is stopped by dilution and the amount of phosphorylase a is then determined as described before [165]. 

Phosphorylase kinase activity has an absolute requirement for Ca +, which binds to the 5-subunit. The amino acid sequence of this subunit is nearly identical to that of calmodulin, with four calcium binding sites, but unlike calmodulin, the 5-subunit is an integral part of the enzyme and does not dissociate from it in the absence of Ca +. In the presence of Ca +, kinase activity is further increased by phosphorylation of the a- and j6-subunits, catalyzed by cAMP-dependent protein kinase and several other kinases. Phosphorylation may activate the enzyme by disinhibiting... 

The answer is b. (Murray, pp 199-207. Scriver, pp 1521-1552. Sack, pp 121-138. Wilson, pp 287-31 77) In the presence of low blood glucose, epinephrine or norepinephrine interacts with specific receptors to stimulate adenylate cyclase production of cyclic AMP Cyclic AMP activates protein kinase, which catalyzes phosphorylation and activation of phosphorylase kinase. Activated phosphorylase kinase activates glycogen phosphorylase, which catalyzes the breakdown of glycogen. Phosphorylase kinase can be activated in two ways. Phosphorylation leads to complete activation of phosphorylase kinase. Alternatively, in muscle, the transient increases in levels of Ca" associated with contraction lead to a partial activation of phosphorylase kinase. Ca" " binds to calmodulin, which is a subunit of phosphorylase kinase. Calmodulin regulates many enzymes in mammalian cells through Ca" binding. 

Pohlig, G. Wingender-Drissen, R. Becker, J.U. Characterization of phosphorylase kinase activities in yeast. Biochem. Biophys. Res. Commun., 114, 331-338 (1983)... 

BVdU is degraded by thymidine phosphorylase more rapidly than the natural substrate, thymidine. This rapid enzymic degradation may present a problem in its clinical use. Moreover, herpes vimses develop resistance to BVdU, apparendy because of mutant vimses that have lower thymidine kinase activity. G. D. Seade has dropped further development of BVdU because of increased animal tumor incidence induced by prolonged dosing (1). 

Stimulation of glycogen breakdown involves consumption of molecules of ATP at three different steps in the hormone-sensitive adenylyl cyclase cascade (Figure 15.19). Note that the cascade mechanism is a means of chemical amplification, because the binding of just a few molecules of epinephrine or glucagon results in the synthesis of many molecules of cyclic / MP, which, through the action of c/ MP-dependent protein kinase, can activate many more molecules of phosphorylase kinase and even more molecules of phosphorylase. For example, an extracellular level of 10 to 10 M epinephrine prompts the for-... 

MP, approximately 30 phosphorylase kinase molecules are activated these in turn activate some 800 molecules of phosphorylase. Each of these catalyzes the formation of many molecules of glucose-l-P. 

A decreased glycolytic rate has been proposed as a cause of muscle fatigue and related to pH inhibition of glycolytic enzymes. Decreasing pH inhibits both phosphorylase kinase and phosphofructokinase (PFK) activities. PFK is rate determining for glycolytic flux and therefore must be precisely matched to the rate of ATP expenditure. The essential characteristic of PFK control is allosteric inhibition by ATP. This inhibition is increased by H and PCr (Storey and Hochachka, 1974 ... 

In hver, one of the serine hydroxyl groups of active phosphorylase a is phosphorylated. It is inactivated by hydrolytic removal of the phosphate by protein phos-phatase-1 to form ph osphorylase h. Reactivation requires rephosphorylation catalyzed by phosphorylase kinase. 

Glycogenolysis increases in muscle several hundred-fold immediately after the onset of contraction. This involves the rapid activation of phosphorylase by activation of phosphorylase kinase by Ca +, the same signal as that which initiates contraction in response to nerve stimulation. Muscle phosphorylase kinase has four... 

Both phosphorylase a and phosphorylase kinase a are dephosphorylated and inactivated by protein phos-phatase-1. Protein phosphatase-1 is inhibited by a protein, inhibitor-1, which is active only after it has been phosphorylated by cAMP-dependent protein kinase. Thus, cAMP controls both the activation and inactivation of phosphorylase (Figure 18-6). Insulin reinforces this effect by inhibiting the activation of phosphorylase b. It does this indirectly by increasing uptake of glucose, leading to increased formation of glucose 6-phosphate, which is an inhibitor of phosphorylase kinase. 

Not only is phosphorylase activated by a rise in concentration of cAMP (via phosphorylase kinase), but glycogen synthase is at the same time converted to the inactive form both effects are mediated via cAMP-dependent protein kinase. Thus, inhibition of glycogenolysis enhances net glycogenesis, and inhibition of glycogenesis enhances net glycogenolysis. Furthermore,... 

NARDINI M, SCACCINI c, PACKER L and VIRGILI F (2000) In vivo inhibition of the activity of phosphorylase kinase, protein kinase C and protein kinase A by caffeic acid and a procyanidin rich pine bark (Pinus maritima) extract Biochimica Biophysica Acta 1474, 219-25. 

SIGNAL INTEGRATION by phosphorylase kinase. Phosphorylase kinase eventually phosphorylates and activates glycogen phosphorylase. Either (or both) phosphorylation and calcium signaling pathways converge at phosphorylase kinase. 

Secondary signals Glucose 6-phosphate activates synthesis. Ca2+-Calmodulin activates degradation by activating phosphorylase kinase. 

Glycogen phosphorylase isoenzymes have been isolated from liver, brain and skeletal muscle. All forms are subject to covalent control with conversion of the inactive forms (GP-b) to the active forms (GP-a) by phosphorylation on specific serine residues. This phosphorylation step, mediated by the enzyme phosphorylase kinase, is initiated by glucagon stimulation of the hepatocyte. Indeed, the same cAMP cascade which inhibits glycogen synthesis simultaneously stimulates glycogenolysis, giving us an excellent example of reciprocal control. 

Calcium ions released from the SR are required to initiate contraction and activate glycogen phosphorylase kinase... 

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