Phosphorylation of phosphorylase kinase

Fig. 7.18. Regulation of glycogen metabolism in muscle. Phosphorylase kinase stands at the center of regulation of glycogen metabolism. Phosphorylase kinase may exist in an active, phosphorylated form and an inactive, unphosphorylated form. Phosphorylation of phosphorylase kinase is triggered by hormonal signals (e.g. adrenahne) and takes place via an activation of protein kinase A in the cAMP pathway. In the absence of hormonal stimulation, phosphorylase kinase can also be activated by an increase in cytosolic Ca. The active phosphorylase kinase stimulates glycogen degradation and inhibits glycogen synthesis, in that, on the one side, it activates glycogen phosphorylase by phosphorylation, and on the other side, it inactivates glycogen synthase by phosphorylation.

Hormonal control of the activity of phosphorylase kinase. Just as the activity of phosphorylase is increased by phosphorylation, so is the activity of its phosphorylase kinase (which may be phosphorylated on two serine residues, one in an a subunit and one in a /3 subunit). Hormonal stimulation (/3-adrenergic) leads to the production of 3, 5 -cyclic AMP ( second messenger ), which stimulates the activity of the cyclic-AMP-dependent protein kinase that catalyzes the phosphorylation of phosphorylase kinase. 

Cyclic AMP triggers a cascade of reactions that ultimately lead to glycogen breakdown. The immediate action of cAMP is to activate a protein kinase that phosphorylates a number of proteins, including phosphorylase kinase. Phosphorylation of phosphorylase kinase converts it from an inactive to an active form, which catalyzes the conversion of phosphorylase b to phosphorylase a (see chapter 9). The cascade of effects triggered by glucagon is shown in figure 12.29. 

Inactivation of the enzyme phosphorylase kinase a is accomplished by dephosphorylation by phosphoprotein phosphatase. A similar phosphoprotein phosphatase is required to convert the active form glycogen phosphorylase a to the inactive glycogen phosphorylase b. It is obvious that a maximal effect of cyclic AMP-stimulated glycogen degradation is obtained when phosphorylation of phosphorylase kinase and glycogen phosphorylase is accompanied by a concomitant... 

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. 

Fig. 7.19. Subunit structure and regulation of phosphorylase kinase of muscle. Phosphorylase kinase is - according to the excitation state of the muscle - regulated by two pathways. On nervous stimulation of the muscle, voltage-controlled Ca channels are opened, the cytosolic Ca concentration increases and Ca binds to calmoduhn, activating phosphorylase kinase. In resting muscle, activation of phosphorylase kinase is triggered by a hormonal signal. A hormonal signal initiates phosphorylation of the a and P subunits of phosphorylase kinase. In the phosphorylated form, Ca binding affinity of the calmodulin subunit (8) is strongly increased and activation is also possible at low Ca concentrations.

The examples of phosphorylase kinase and protein phosphatase I illustrate some important principles of regulation of enzyme activity by phosphorylation and dephosphorylation events. They clearly indicate how different signal transduction paths can meet in key reactions of metabolism, how signals can be coordinated with one another and how common components of a regulation network can be activated by different signals. The following principles are highlighted ... 

FIGURE 6-31 Regulation of glycogen phosphorylase activity by covalent modification. In the more active form of the enzyme, phosphorylase a, specific Ser residues, one on each subunit, are phosphorylated. Phosphorylase a is converted to the less active phosphorylase b by enzymatic loss of these phosphoryl groups, promoted by phosphorylase phosphatase. Phosphorylase b can be reconverted (reactivated) to phosphorylase a by the action of phosphorylase kinase. 

Activation of phosphorylase kinase Phosphorylase kinase exists in two forms an inactive "b" form and an active "a" form. Active cAMP-dependent protein kinase phosphorylates the inactive form of phosphorylase kinase, resulting in its activation (see Figure... 

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. 

GDP production and the inactivation of the G protein. If hormone remains bound, the G protein can be activated again. Meanwhile, the cAMP produced binds to the R subunits of cAMP-dependent protein kinase, leading to the dissociation of two catalytic subunits, which can then phosphorylate critical proteins in this case the activation of phosphorylase kinase is shown. 

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.

Glycogen degradation requires phosphorylated (active) phosphorylase kinase to maintain the production of phosphorylated (active) phosphorylase a the a-l,6-glucosidase is required to remove branch molecules from partially degraded glycogen, and phosphorylated (active) inhibitor protein is required to inactivate phosphoprotein phosphatase. 

Figure 21.13. Activation of Phosphorylase Kinase. Phosphorylase kinase is activated by hormones that lead to the phosphorylation of the P subunit and by Ca2+ binding of the 8 subunit. Both types of stimulation are required for maximal enzyme activity. 

Protein kinase A phosphorylates the P subunit of phosphorylase kinase, which subsequently activates glycogen phosphorylase. 

Multiple phosphorylation. Protein kinase A activates muscle phosphorylase kinase by rapidly phosphorylating its P subunits. The a subunits of phosphorylase kinase are then slowly phosphorylated, which makes the a and P subunits susceptible to the action of protein phosphatase 1. What is the functional significance of the slow phosphorylation of a ... 

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-... 

B. Under these conditions, cAMP levels would remain elevated. Phosphorylation of pyruvate kinase causes its inactivation. Phosphorylase kinase and phosphorylase are activated by phosphorylation. Protein kinase A is not regulated by phosphorylation. 

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