15.3 Coordinated Regulation of Glycogen Breakdown and Synthesis in Chapter 15 The Metabolism of Glycogen in Animals

15.3 Coordinated Regulation of Glycogen Breakdown and Synthesis

As we have seen, the mobilization of stored glycogen is brought about by glycogen phosphorylase, which degrades glycogen to glucose 1-phosphate (Fig. 15-3). Glycogen phosphorylase provides an especially instructive case of enzyme regulation. It was one of the first known examples of an allosterically regulated enzyme and the first enzyme shown to be controlled by reversible phosphorylation. It was also one of the first allosteric enzymes for which the detailed three-dimensional structures of the active and inactive forms were revealed by x-ray crystallographic studies. Glycogen phosphorylase also illustrates how isozymes play their tissue-specific roles.

Glycogen Phosphorylase Is Regulated by Hormone-Stimulated Phosphorylation and by Allosteric Effectors

A signed photo of Earl W. Sutherland. — Earl W. Sutherland, Jr., 1915–1974

In the late 1930s, Carl and Gerty Cori (Box 15-1) discovered that the glycogen phosphorylase of skeletal muscle exists in two interconvertible forms: glycogen phosphorylase a, which is catalytically active, and glycogen phosphorylase b, which is much less active. (Note that glycogen phosphorylase is often referred to simply as phosphorylase — so honored because it was the first phosphorylase to be discovered; the shortened name has persisted in common usage and in the literature.) Subsequent studies by Earl Sutherland showed that phosphorylase b predominates in resting muscle, but during vigorous muscular activity, epinephrine triggers phosphorylation of phosphorylase b, converting it to its more active form, phosphorylase a (Fig. 15-11). In the liver, glucagon triggers phosphorylation of phosphorylase b, converting it to its active form.

A figure shows how muscle glycogen phosphorylase can be regulated using covalent modification. — FIGURE 15-11 Regulation of muscle glycogen phosphorylase by covalent modification. In the more active form of the enzyme, phosphorylase a, ${Ser}^{14}$ $Ser Superscript 14$ residues, one on each subunit, are phosphorylated. Phosphorylase a is converted to the less active form, phosphorylase b, by enzymatic loss of these phosphoryl groups, catalyzed by phosphoprotein phosphatase 1 (PP1). Phosphorylase b can be reconverted (reactivated) to phosphorylase a by the action of phosphorylase b kinase.

FIGURE 15-11 Regulation of muscle glycogen phosphorylase by covalent modification. In the more active form of the enzyme, phosphorylase a, ${Ser}^{14}$ $Ser Superscript 14$ residues, one on each subunit, are phosphorylated. Phosphorylase a is converted to the less active form, phosphorylase b, by enzymatic loss of these phosphoryl groups, catalyzed by phosphoprotein phosphatase 1 (PP1). Phosphorylase b can be reconverted (reactivated) to phosphorylase a by the action of phosphorylase b kinase.

A blue rectangle is divided into two halves and labeled phosphorylase italicized b end italics light red highlighted (less active). The left half has a half circle at the bottom, and the right half has a half circle at the top. The left side has S e r superscript 14 at the top center with a bond extending up above it to C H 2 further bonded to O H. The right half has S e r superscript 14 at the bottom center with a bond extending down below it to C H 2 further bonded to O H. A curved arrow extending right is labeled phosphorylase italicized b end italics. A green circle around a green triangle is above the start of the arrow and a dashed arrow points to it from text reading, glucagon (liver). A second green circle around a green triangle is at the right end of the arrow and a dashed arrow points from it to text reading, epinephrine, up arrow symbol [C a 2 plus], up arrow symbol [A M P] (muscle). A curved arrow meets it from below to show that 2 A T P are added and 2 A D P leave. This yields a similar blue structure with its top side moved to the left of the bottom side, meaning that the left and right sides are parallel diagonal lines that run from upper left to lower right. There are short lines radiating from all sides of the structure, which is labeled phosphorylase italicized a end italics red highlighted (active). The left half still has a half circle at the bottom and the right half still has a half circle at the top. The left side has S e r superscript 14 above the top left side with a bond extending down to C H 2 within the blue structure, where it is further bonded to O bonded to a green circle labeled P. The right side has S e r superscript 14 below the bottom right side with a bond extending up to C H 2 within the blue structure, where it is further bonded to O bonded to a green circle labeled P.

Sutherland discovered the second messenger cAMP, which increases in concentration in response to stimulation by epinephrine (in muscle; see Fig. 12-4) or glucagon (in liver). Elevated [cAMP] initiates an enzyme cascade (Fig. 15-12), in which a catalyst activates a catalyst, which activates a catalyst. As we saw in Chapter 12, such cascades allow exponential amplification of the initial signal. The rise in [cAMP] activates cAMP-dependent protein kinase, also called protein kinase A (PKA). PKA then phosphorylates and activates phosphorylase b kinase, which catalyzes the phosphorylation of glycogen phosphorylase b, activating it and thus stimulating glycogen breakdown.

A figure shows the cascade mechanism of action of epinephrine in a myocyte and of glucagon in a hepatocyte. — FIGURE 15-12 Cascade mechanism of epinephrine and glucagon action. By binding to specific surface receptors, either epinephrine acting on a myocyte (left) or glucagon acting on a hepatocyte (right) activates a GTP-binding protein, $G_{s α}$ $upper G Subscript s alpha$ . Active $G_{s α}$ $upper G Subscript s alpha$ triggers a rise in [cAMP], activating PKA. This sets off a cascade of phosphorylations; PKA activates phosphorylase b kinase, which then activates glycogen phosphorylase. Such cascades effect a large amplification of the initial signal; the figures in pink boxes are certainly low estimates of the actual increase in number of molecules at each stage of the cascade. The resulting breakdown of glycogen provides glucose, which in the myocyte can supply ATP (via glycolysis) for muscle contraction and in the hepatocyte is released into the blood to counter the low blood glucose.

FIGURE 15-12 Cascade mechanism of epinephrine and glucagon action. By binding to specific surface receptors, either epinephrine acting on a myocyte (left) or glucagon acting on a hepatocyte (right) activates a GTP-binding protein, $G_{s α}$ $upper G Subscript s alpha$ . Active $G_{s α}$ $upper G Subscript s alpha$ triggers a rise in [cAMP], activating PKA. This sets off a cascade of phosphorylations; PKA activates phosphorylase b kinase, which then activates glycogen phosphorylase. Such cascades effect a large amplification of the initial signal; the figures in pink boxes are certainly low estimates of the actual increase in number of molecules at each stage of the cascade. The resulting breakdown of glycogen provides glucose, which in the myocyte can supply ATP (via glycolysis) for muscle contraction and in the hepatocyte is released into the blood to counter the low blood glucose.

The figure is divided into two halves with the left half labeled myocyte and the right half labeled hepatocyte. A rectangle labeled italicized x end italics molecules is at the top center, overlapping a dashed line that separates the halves. A lighter rectangle is in the center of the illustration, overlapping each half. The top left border of the lighter rectangle contains an orange rectangle with a square cutout in its upper side, outside of the lighter rectangle. A green square labeled epinephrine is shown above the square cutout. A dashed arrow points from the bottom of the orange rectangle beneath epinephrine to a green circle containing a green triangle next to G subscript s Greek letter alpha. The top right border of the lighter rectangle contains an orange circle with a triangular cutout in its upper side, outside of the lighter rectangle. A blue triangle labeled glucagon is shown above the triangular cutout. A dashed arrow points from the bottom of the orange rectangle beneath glucagon to a green circle containing a green triangle next to G subscript s Greek letter alpha. A dashed arrow points down from G subscript s Greek letter alpha to a green circle containing a green triangle above an arrow labeled adenylyl cyclase pointing from A T P on the left to cyclic A M P on the right. A rectangle nearby reads, 20 x molecules. A curved arrow points down from cyclic A M P to an arrow from inactive P K A on the left to active P K A on the right. A rectangle nearby reads, 10 x molecules. A dashed curved arrow points down from active P K A to a green circle containing a green triangle above an arrow pointing from inactive phosphorylase italicized b end italics kinase on the left to active phosphorylase italicized b end italics kinase on the right. A rectangle nearby reads, 100 x molecules. A dashed arrow from upward arrow symbol [C a 2 plus] in the myocyte outside of the lighter rectangle points across its boundary to the green circle containing a green triangle above the arrow pointing from inactive phosphorylase italicized b end italics kinase on the left to active phosphorylase italicized b end italics kinase on the right. A dashed arrow points down from active phosphorylase italicized b end italics kinase to a green circle containing a green triangle above an arrow pointing from inactive glycogen phosphorylase italicized b end italics on the left to active glycogen phosphorylase italicized a end italics on the right. A rectangle nearby reads, 1,000 x molecules. A dashed arrow points down from active glycogen phosphorylase italicized a end italics to a green circle containing a green triangle above an arrow pointing from glycogen on the left to glucose 1-phosphate on the right. A rectangle nearby reads, 10,000 molecules. A dashed arrow from upward arrow symbol [A M P] in the myocyte outside of the lighter rectangle points across its boundary to the green circle containing a green triangle to the left of the arrow from active glycogen phosphorylase italicized a end italics to the green circle containing a green triangle above the arrow from glycogen to glucose 1-phosphate. Two arrows point down from glucose 1-phosphate. The left-hand arrow runs horizontally left, then down out of the lighter rectangle to end at glycolysis in the myocyte, from which an arrow points down to muscle contraction in the myocyte. The right-hand arrow points to a second arrow that runs out of the lighter rectangle into the hepatocyte to end at glucose, from which a second arrow points to blood glucose below. A nearby rectangle reads, 10,000 x molecules.

In muscle, this provides fuel for glycolysis to sustain muscle contraction for the fight-or-flight response signaled by epinephrine. In liver, glycogen breakdown counters the low blood glucose signaled by glucagon, by releasing glucose into the blood. These different roles are reflected in subtle differences in the regulatory mechanisms in muscle and liver. The glycogen phosphorylases of liver and muscle are isozymes, encoded by different genes and differing in their regulatory properties.

Figure 15-12 shows the pathway for hormonal regulation of glycogen phosphorylase activity: epinephrine or glucagon initiates the cascade that activates phosphorylase b kinase. Phosphorylase b kinase activates phosphorylase by transferring a phosphoryl group to ${Ser}^{14}$ $Ser Superscript 14$ on each of the two identical subunits of phosphorylase b, triggering a conformation change from the T state (phosphorylase b) to the R state (phosphorylase a), shown in detail in Figure 6-40. Superimposed on the hormonal activation of phosphorylase b kinase are allosteric control mechanisms (Fig. 15-12). In muscle, $Ca Superscript 2 plus$ $Ca Superscript 2 plus$ , the signal for muscle contraction, binds to and activates phosphorylase b kinase. ${Ca}^{2 +}$ $Ca Superscript 2 plus$ binds to phosphorylase b kinase through its δ subunit, which is calmodulin (see Fig. 12-17). AMP, which accumulates in vigorously contracting muscle as a result of ATP breakdown, binds to and activates phosphorylase, speeding the release of glucose 1-phosphate from glycogen. When ATP levels are adequate, ATP blocks the allosteric site to which AMP binds (see Fig. 6-40), inactivating phosphorylase.

When the muscle returns to rest, a second enzyme, phosphoprotein phosphatase 1 (PP1), removes the phosphoryl groups from phosphorylase a, converting it to the less active form, phosphorylase b (see Fig. 15-11).

Like the enzyme of muscle, the glycogen phosphorylase of liver is regulated hormonally (by phosphorylation/dephosphorylation) and allosterically. The dephosphorylated form is essentially inactive. When the blood glucose level is too low, glucagon (acting through the cascade mechanism shown in Fig. 15-12) activates phosphorylase b kinase, which in turn converts phosphorylase b to its active a form, initiating the release of glucose into the blood. When blood glucose levels return to normal, glucose enters hepatocytes and binds to an inhibitory allosteric site on phosphorylase a. This binding also produces a conformational change that exposes the phosphorylated Ser residues to PP1, which catalyzes their dephosphorylation and inactivates the phosphorylase (Fig. 15-13). The allosteric site for glucose allows liver glycogen phosphorylase to act as its own glucose sensor and to respond appropriately to changes in blood glucose.

A figure shows how glycogen phosphorylase of the liver can be used as a glucose sensor. — FIGURE 15-13 Glycogen phosphorylase of liver as a glucose sensor. Glucose binding to an allosteric site of the phosphorylase a isozyme of liver induces a conformational change that exposes its phosphorylated Ser residues to the action of phosphoprotein phosphatase 1 (PP1). This phosphatase converts phosphorylase a to phosphorylase b, sharply reducing the activity of phosphorylase and slowing glycogen breakdown in response to high blood glucose. Insulin also acts indirectly to stimulate PP1 and slow glycogen breakdown.

A blue parallelogram is divided into two halves and labeled phosphorylase italicized a end italics light red highlighted (active active). There are short lines radiating from all sides of the structure. The left half has a half circle at the bottom, and the right half has a half circle at the top. These half circles are labeled allosteric site empty. The left side has a bond extending down from the upper left side to C H 2 further bonded to O bonded to a green circle labeled P. The right side has a bond extending up from the lower right side to C H 2 further bonded to O bonded to a green circle labeled P. An arrow pointing right is joined by an arrow showing the addition of 2 glucose. This yields a similar structure labeled phosphorylase italicized a end italics that now has its top and bottom sides lined up to form a rectangle. It still has narrow lines radiating from all sides. The half-circles are now colored red and labeled G l c. There is a bond from the top center of the left side to C H 2 bonded to O bonded to a green circle labeled P, forming a vertical line up from the rectangle. There is a bond from the bottom center of the right side to C H 2 bonded to O bonded to a green circle labeled P, forming a vertical line down from the rectangle. An arrow pointing right is labeled P P 1. A dashed arrow points down from insulin to a green circle containing a green triangle on the arrow. An arrow curves away to show the loss of 2 P subscript i. This yields a product labeled phosphorylase italicized b end italics red highlighted (less active). It is similar except that there are no short lines from radiating from all sides, the C H 2 bonded to the top center left is now bonded to O H, and the C H 2 bonded to the bottom center right is now bonded to O H.

Glycogen Synthase Also Is Subject to Multiple Levels of Regulation

Like glycogen phosphorylase, glycogen synthase can exist in phosphorylated and unphosphorylated forms (Fig. 15-14). Its active form, glycogen synthase a, is unphosphorylated. Glycogen synthase kinase 3 (GSK3) adds phosphoryl groups to three Ser residues near the carboxyl terminus of glycogen synthase a converting it to glycogen synthase b, which is inactive unless its allosteric activator, glucose 6-phosphate, is present. The action of GSK3 is hierarchical; it cannot phosphorylate glycogen synthase until another protein kinase, casein kinase II (CKII), has first phosphorylated the glycogen synthase on a nearby residue, an event called priming (Fig. 15-15a). AMP-activated protein kinase (AMPK), which associates with glycogen granules through its carbohydrate-binding domain, also phosphorylates glycogen synthase, inhibiting glycogen synthesis during periods of metabolic stress, signaled by high [AMP] and low [ATP].

A figure shows how G S K 3 affects the activity of glycogen synthase and compares the structures of glycogen synthase italicized a end italics and glycogen synthase italicized b end italics. — FIGURE 15-14 Effects of GSK3 on glycogen synthase activity. Glycogen synthase a, the active form, has three Ser residues near its carboxyl terminus. Their phosphorylation by glycogen synthase kinase 3 (GSK3) converts glycogen synthase to its inactive b form. Insulin favors the active a form of glycogen synthase by blocking the activity of GSK3 and activating phosphoprotein phosphatase 1 (PP1). In muscle, epinephrine activates PKA, which phosphorylates the glycogen-targeting protein Gm on a site that causes dissociation of PP1 from glycogen. Glucose 6-phosphate favors dephosphorylation of glycogen synthase by binding to it and promoting a conformation that is a good substrate for PP1.

A purple oval labeled glycogen synthase italicized a end italics active is shown at the nine o’clock position of a circle with glycogen synthase italicized b end italics inactive at the three o’clock position. A curved arrow across the top of the circle points from glycogen synthase italicized a end italics to glycogen synthase italicized b end italics and a second curved arrow across the bottom of the circle points from glycogen synthase italicized b end italics to glycogen synthase italicized a end italics. Glycogen synthase italicized a end italics has three bonds to O H at its upper left side and these are labeled, serines near carboxyl terminus. Near the beginning of the curved upward arrow, a smaller curved arrow shows that A T P comes in, then meets the larger curved arrow beneath a yellow oval labeled C K Roman numeral 2, then shows that A D P leaves. At the 12 o’clock position, a yellow rectangle labeled G S K 3 is beneath the top of the curved arrow. A curved arrow above it shows that 3 A T P comes in from the left and 3 A D P leaves to the right. A light yellow box labeled insulin above has a dashed arrow pointing down to a red circle containing a red X directly above G S K 3 in the center of the arrow between A T P and A D P. Glycogen synthase italicized G end italics has three red circles labeled P bonded to its upper right side and these are labeled, phosphoserines near carboxyl terminus. At the 6 o’clock position of the curved circle pointing back to glycogen synthase italicized a end italics, there is a light green circle labeled P P 1 just above the arrow. Clockwise from just to its right, four arrows point to the large bottom curved arrow as follows: A yellow box labeled glucose has a dashed arrow pointing to a green circle containing a green triangle; a yellow box labeled glucose 6-phosphate has a dashed arrow pointing to a green circle containing a green triangle; a yellow box labeled insulin has a dashed arrow pointing to a green circle containing a green triangle; and a yellow box labeled glucagon, epinephrine has a dashed arrow pointing to a red circle labeled with a red X. An arrow curves away to show the loss of 3 P subscript i before the arrow reaches glycogen synthase italicized a end italics again.

A two-part figure shows the priming of G S K 3 phosphorylation of glycogen synthase by showing the association between glycogen synthase kinase 3 and its substrate in part a and then showing the effects of phosphorylation in G S K 3, which leads to formation of a pseudosubstrate region, in part b. — FIGURE 15-15 Priming of GSK3 phosphorylation of glycogen synthase. (a) Glycogen synthase kinase 3 first associates with its substrate (glycogen synthase) by interaction between three positively charged residues ${(Arg}^{96}, {Arg}^{180}, {Lys}^{205})$ $left-parenthesis Arg Superscript 96 Baseline comma Arg Superscript 180 Baseline comma Lys Superscript 205 Baseline right-parenthesis$ and a phosphoserine residue at position $+ 4$ $plus 4$ in the substrate. (For orientation, the Ser or Thr residue to be phosphorylated in the substrate is assigned the index 0. Residues on the amino-terminal side of this residue are numbered $- 1, - 2,$ $negative 1 comma negative 2 comma$ and so forth; residues on the carboxyl-terminal side are numbered $+ 1, + 2,$ $plus 1 comma plus 2 comma$ and so forth.) This association aligns the active site of the enzyme with a Ser residue at position 0, which it phosphorylates. This creates a new priming site, and the enzyme moves down the protein to phosphorylate the Ser residue at position $negative 4$ $negative 4$ , and then the Ser at $negative 8$ $negative 8$ . (b) GSK3 has a Ser residue near its amino terminus that can be phosphorylated by PKA or PKB. This produces a “pseudosubstrate” region in GSK3 that folds into the priming site and makes the active site inaccessible to another protein substrate, inhibiting GSK3 until the priming phosphoryl group of its pseudosubstrate region is removed by PP1. Other proteins that are substrates for GSK3 also have a priming site at position $plus 4$ $plus 4$ , which must be phosphorylated by another protein kinase before GSK3 can act on them.

FIGURE 15-15 Priming of GSK3 phosphorylation of glycogen synthase. (a) Glycogen synthase kinase 3 first associates with its substrate (glycogen synthase) by interaction between three positively charged residues ${(Arg}^{96}, {Arg}^{180}, {Lys}^{205})$ $left-parenthesis Arg Superscript 96 Baseline comma Arg Superscript 180 Baseline comma Lys Superscript 205 Baseline right-parenthesis$ and a phosphoserine residue at position $+ 4$ $plus 4$ in the substrate. (For orientation, the Ser or Thr residue to be phosphorylated in the substrate is assigned the index 0. Residues on the amino-terminal side of this residue are numbered $- 1, - 2,$ $negative 1 comma negative 2 comma$ and so forth; residues on the carboxyl-terminal side are numbered $+ 1, + 2,$ $plus 1 comma plus 2 comma$ and so forth.) This association aligns the active site of the enzyme with a Ser residue at position 0, which it phosphorylates. This creates a new priming site, and the enzyme moves down the protein to phosphorylate the Ser residue at position $negative 4$ $negative 4$ , and then the Ser at $negative 8$ $negative 8$ . (b) GSK3 has a Ser residue near its amino terminus that can be phosphorylated by PKA or PKB. This produces a “pseudosubstrate” region in GSK3 that folds into the priming site and makes the active site inaccessible to another protein substrate, inhibiting GSK3 until the priming phosphoryl group of its pseudosubstrate region is removed by PP1. Other proteins that are substrates for GSK3 also have a priming site at position $plus 4$ $plus 4$ , which must be phosphorylated by another protein kinase before GSK3 can act on them.

Part a shows a bar labeled glycogen synthase that has the following structure: three dashes, A, highlighted S bonded to O H above, V, P, P, highlighted S bonded to O H above with negative 8 written below, P, S, L, highlighted S bonded to highlighted phosphate above and with negative 4 written below, R, H, S, highlighted S bonded to O H above and with 0 written below, P, H, Q, highlighted S bonded to O H above and with plus 4 written below, E, D, E, E, three dashes. Lines pointing to negative 8, negative 4, and 0 read, S e r residues phosphorylated in glycogen synthase. A yellow rectangular structure labeled G S K is positioned over the right side of glycogen synthase and has two circular cutouts below and a horizontal piece extending to its lower right. The left-hand opening is between its narrow left side, which curves slightly right to end at the space between H and S at positions negative 2 and negative 3, and a narrow central piece, which ends above H at position positive 2. The left-hand cutout has short radiating lines on all sides, A T P in its center, and O H from highlighted S at position 0 extending up into it. The top of the cutout is labeled active site. The right-hand opening contains a highlighted phosphate group bonded to S at position plus 4. S is bonded to highlighted O bonded to highlighted P bonded to highlighted O minus to the left and above and double bonded to O to the right. Accompanying text reads, priming site phosphorylated by casein kinase Roman numeral 2. Around the top of the right hand side, there are three circles labeled as follows: A r g superscript 96 at the upper left, A r g superscript 180 at the center, and L y s superscript 205 at the upper right. An arrow points left from G S K 3. A dashed outline of G S K 3 is shown to its left with an arrow pointing further left. This dashed outline has its left side contacting glycogen synthase above S at position negative 6 and its middle piece above R and H at positions negative 3 to negative 2. Part b shows G S K 3 with the horizontal piece that had extended to the right now folded underneath to run horizontally toward the left. It has N H 3 plus at its left side, then three dashes, then R, P above 0, R, T, T, highlighted S bonded to highlighted phosphate above in the right-hand cutout above position plus 4, F, then a curve up the right side with A, E, S, C, then three dashes.

Insulin favors activation of glycogen synthase by blocking the activity of GSK3 and activating PP1. In muscle, epinephrine activates PKA, which phosphorylates the glycogen-targeting protein Gm (see Fig. 15-16), a regulatory subunit of PP1 in muscle, on a site that causes dissociation of PP1 from glycogen, effectively blocking its action on glycogen synthase.

In liver, conversion of glycogen synthase b to the active form is promoted by PP1, which is bound to the glycogen particle by its regulatory subunit in liver, Gl. PP1 removes the phosphoryl groups from the three Ser residues phosphorylated by GSK3. Glucose 6-phosphate binds to an allosteric site on glycogen synthase, making the enzyme a better substrate for dephosphorylation by PP1 and causing its activation. By analogy with glycogen phosphorylase, which acts as a glucose sensor, glycogen synthase can be regarded as a glucose 6-phosphate sensor. In muscle, a different phosphatase may have the role played by PP1 in liver, activating glycogen synthase by dephosphorylating it.

As we saw in Chapter 12, one way in which insulin triggers intracellular changes is by activating a protein kinase (PKB) that, in turn, phosphorylates and inactivates GSK3 (see Fig. 12-23). Phosphorylation of a Ser residue near the amino terminus of GSK3 converts that region of the protein to a pseudosubstrate, which folds into the site at which the priming phosphorylated Ser residue normally binds (Fig. 15-15b). This prevents GSK3 from binding the priming site of a real substrate, thereby inactivating the enzyme and tipping the balance in favor of dephosphorylation of glycogen synthase by PP1. Glycogen phosphorylase can also affect the phosphorylation of glycogen synthase: active glycogen phosphorylase directly inhibits PP1, preventing it from activating glycogen synthase (Fig. 15-14).

Insulin stimulates glycogen synthesis by activating PP1 and by inactivating GSK3. This single enzyme, PP1, can remove phosphoryl groups from all three of the enzymes phosphorylated in response to glucagon (liver) and epinephrine (liver and muscle): phosphorylase kinase, glycogen phosphorylase, and glycogen synthase. The catalytic subunit of PP1 (PP1c) does not exist free in the cytosol, but is tightly bound to its target proteins through a tissue-specific regulatory subunit, one of a family of glycogen-targeting proteins that bind glycogen and each of the three enzymes (Fig. 15-16). PP1 is itself subject to covalent and allosteric regulation: it is inactivated when phosphorylated by PKA and is allosterically activated by glucose 6-phosphate.

A figure shows the function of the glycogen-targeting protein G subscript M in two steps. — FIGURE 15-16 Glycogen-targeting protein Gm. Gm is a regulatory subunit of PP1 in muscle, and one of a family of glycogen-targeting proteins that serve as a scaffold, binding other proteins (including PP1) to glycogen particles. Gm can be phosphorylated at two different sites in response to insulin or epinephrine. Insulin-stimulated phosphorylation of Gm site 1 activates PP1, which dephosphorylates phosphorylase kinase, glycogen phosphorylase, and glycogen synthase. Epinephrine-stimulated phosphorylation of Gm site 2 by PKA causes dissociation of PP1 from the glycogen particle, preventing its access to glycogen phosphorylase and glycogen synthase. PKA also phosphorylates a protein (inhibitor 1) that, when phosphorylated, inhibits PP1. By these means, insulin stimulates glycogen synthesis and inhibits glycogen breakdown, whereas epinephrine (or glucagon in the liver) has the opposite effects.

FIGURE 15-16 Glycogen-targeting protein Gm. Gm is a regulatory subunit of PP1 in muscle, and one of a family of glycogen-targeting proteins that serve as a scaffold, binding other proteins (including PP1) to glycogen particles. Gm can be phosphorylated at two different sites in response to insulin or epinephrine. Insulin-stimulated phosphorylation of Gm site 1 activates PP1, which dephosphorylates phosphorylase kinase, glycogen phosphorylase, and glycogen synthase. Epinephrine-stimulated phosphorylation of Gm site 2 by PKA causes dissociation of PP1 from the glycogen particle, preventing its access to glycogen phosphorylase and glycogen synthase. PKA also phosphorylates a protein (inhibitor 1) that, when phosphorylated, inhibits PP1. By these means, insulin stimulates glycogen synthesis and inhibits glycogen breakdown, whereas epinephrine (or glucagon in the liver) has the opposite effects.

Within a glycogen granule, a structure has a light blue oval labeled glycogen phosphorylase bonded to a circle labeled P below with a green circle labeled P P 1 to its upper right above a tilted blue oval to its lower right labeled phosphorylase kinase that runs slightly behind the lower right end of glycogen phosphorylase before extending down slightly to the lower right with a circle labeled P connected below. To the right of the bottom of P P 1 and the top of phosphorylase kinase, horizontally aligned with glycogen phosphorylase, there is a purple oval labeled glycogen synthase bonded to a circle labeled P below. Behind this structure, a slightly bent horizontal structure labeled G subscript M runs from just after the start of glycogen phosphorylase to just before the end of glycogen synthase. An arrow numbered 1 and insulin-sensitive kinase points to a similar structure to the right. Insulin has a dashed arrow pointing down to a green circle containing a green triangle just above the arrow showing step 1. This yields a similar structure to the starting structure except that glycogen phosphorylase, phosphorylase kinase, and glycogen synthase are no longer bonded to circles labeled P. P P 1 is surrounded by radiating lines and G subscript M is bonded to a circle labeled P above and to the right of P P 1. Text below reads, upward arrow symbol glycogen synthesis; downward arrow symbol glycogen breakdown. An arrow pointing right is labeled 2 and P K A. Epinephrine has a dashed arrow pointing down to a green circle containing a green triangle above this arrow. A small arrow points to a crescent with radiating lines all around labeled inhibitor 1 from which an arrow points to a similar shape with no radiating lines and a circle labeled P bound to its lower right side. A dashed arrow labeled phosphorylated inhibitor 1 binds and inactivates P P 1 points from this crescent to a similar crescent that fits against the right side of P P 1. The arrow labeled 2 also has a branch that splits in two. The top half points to the green sphere labeled P P 1 bound to the crescent bonded to a circle labeled P. The bottom half points to a structure with a light blue oval labeled glycogen phosphorylase with an open space to its upper right where P P 1 had been and a blue oval representing phosphorylase kinase to its right. A purple oval labeled glycogen synthase is to the right of these, aligned horizontally with glycogen phosphorylase. The large bent purple horizontal piece is still present behind these, but now is bonded to circles labeled P on either side of the cavity that had contained the green circle labeled P P 1. Text below reads, downward arrow symbol glycogen synthesis; upward arrow symbol glycogen breakdown.

Allosteric and Hormonal Signals Coordinate Carbohydrate Metabolism Globally

Having looked at the mechanisms that regulate individual enzymes, we can now consider the overall shifts in carbohydrate metabolism that occur in the well-fed state, during fasting, and in the fight-or-flight response — signaled by insulin, glucagon, and epinephrine, respectively. We need to contrast two cases in which regulation serves different ends: (1) the role of hepatocytes in supplying glucose to the blood; and (2) the selfish use of carbohydrate fuels by extrahepatic tissues, typified by skeletal muscle (myocytes), to support their own activities.

After ingestion of a carbohydrate-rich meal, the elevation of blood glucose triggers insulin release (Fig. 15-17, top). In a hepatocyte, insulin has two immediate effects: it inactivates GSK3; and it activates a protein phosphatase, probably PP1. These two actions fully activate glycogen synthase. PP1 also inactivates glycogen phosphorylase a and phosphorylase kinase by dephosphorylating both, effectively stopping glycogen breakdown. Glucose enters the hepatocyte through the high-capacity transporter GLUT2, always present in the plasma membrane, and the elevated intracellular glucose leads to dissociation of hexokinase IV (glucokinase) from its nuclear regulatory protein (Fig. 14-21). Hexokinase IV enters the cytosol and phosphorylates glucose, stimulating glycolysis and supplying the precursor for glycogen synthesis. Under these conditions, hepatocytes use the excess glucose in the blood to synthesize glycogen, up to the limit of about 10% of the total weight of the liver.

A figure shows the complex interactions that affect the regulation of carbohydrate metabolism in the liver. — FIGURE 15-17 Regulation of carbohydrate metabolism in the liver. Colored arrows indicate causal relationships between the changes they connect. For example, an arrow from $↓ A$ $down-arrow upper A$ to $↑ B$ $up-arrow upper B$ means that a decrease in A causes an increase in B. Red arrows connect events that result from high blood glucose; blue arrows connect events that result from low blood glucose.

FIGURE 15-17 Regulation of carbohydrate metabolism in the liver. Colored arrows indicate causal relationships between the changes they connect. For example, an arrow from $↓ A$ $down-arrow upper A$ to $↑ B$ $up-arrow upper B$ means that a decrease in A causes an increase in B. Red arrows connect events that result from high blood glucose; blue arrows connect events that result from low blood glucose.

The top half of the figure consists of red arrows ending at red boxes, and the bottom half of the figure consists of blue arrows ending at blue boxes. The top of the figure begins with high blood glucose, from which a red arrow extends downward to the left and right. The left-hand arrow points to upward arrow insulin, from which three red arrows point down. The first short red arrow from upward arrow insulin points to upward arrow insulin-sensitive protein kinase, from which a red arrow points to upward arrow P P 1, from which three more red arrows point downward. The left-hand red arrow points down to downward arrow phosphorylase kinase, from which a second red arrow points to downward arrow glycogen phosphorylase. The middle red arrow from upward arrow P P 1 also points to downward arrow glycogen phosphorylase, from which another red arrow points to a red box labeled downward arrow glycogen breakdown. The right-hand red arrow from upward arrow P P 1 points to upward arrow glycogen synthase, which is also in the pathway from the middle arrow from upward arrow insulin. The middle arrow from upward arrow insulin has a short red arrow to upward arrow P K B, which has a red arrow to downward arrow G S K - 3, from which another red arrow points to upward arrow glycogen synthase. A red arrow points from upward arrow glycogen synthase to a red box labeled upward arrow glycogen synthesis. The right-hand arrow from upward arrow insulin points to synthesis of hexokinase Roman numeral 2, P F K - 1, pyruvate kinase, from which a red arrow points to a red box labeled upward arrow glycolysis. The right-hand arrow from high blood glucose at the very top runs through G L U T 2 to point to upward arrow [glucose] subscript inside, from which a red arrow points down and splits into a left-hand piece that ends at the red box labeled upward arrow glycogen synthesis and a right-hand piece that ends at the light red box labeled upward arrow glycolysis. At the very bottom, low blood glucose has a blue arrow pointing to upward arrow glucagon, which has a blue arrow to upward arrow [c A M P], which has a blue arrow pointing to upward arrow P K A, from which there are four blue arrows. The left-hand blue arrow points to upward arrow phosphorylase kinase, from which a blue arrow points to upward arrow glycogen phosphorylase, from which a short blue arrow points to a blue box labeled upward arrow glycogen breakdown beneath the red box labeled downward arrow glycogen breakdown. The second blue arrow from upward arrow P K A points to downward arrow glycogen synthase, from which a short blue arrow points to a blue box labeled downward arrow glycogen synthesis beneath the red box labeled upward arrow glycogen synthesis. The third blue arrow from upward arrow P K A is short and points to upward arrow F B Pase - 2, downward arrow P F K - 2, from which a blue arrow points up, jags to the right, then continues up to downward arrow [F 26 B P], from which a short blue arrow points up to downward arrow P F K - 1, from which a short blue arrow points to a blue box labeled downward arrow glycolysis beneath the red box labeled upward arrow glycolysis. The fourth blue arrow from upward arrow P K A points to downward arrow pyruvate kinase L, from which a long blue arrow points to a blue box labeled downward arrow glycolysis beneath the red box labeled upward arrow glycolysis.

Between meals, or during an extended fast, the blood glucose level drops, triggering the release of glucagon, which, acting through the cascade shown in Figure 15-12, activates PKA. PKA mediates all the effects of glucagon (Fig. 15-17, bottom). It phosphorylates phosphorylase kinase, activating it and leading to the activation of glycogen phosphorylase. It phosphorylates glycogen synthase, inactivating it and blocking glycogen synthesis. It phosphorylates PFK-2/FBPase-2, leading to a drop in the concentration of the regulator fructose 2,6-bisphosphate, which has the effect of inactivating the glycolytic enzyme PFK-1 and activating the gluconeogenic enzyme FBPase-1 (see Fig. 14-24). And it phosphorylates and inactivates the glycolytic enzyme pyruvate kinase. Under these conditions, the liver produces glucose 6-phosphate by glycogen breakdown and by gluconeogenesis, and it stops using glucose to fuel glycolysis or make glycogen, maximizing the amount of glucose it can release to the blood. This release of glucose is possible only in liver and kidney, because other tissues lack glucose 6-phosphatase (Fig. 15-6).

The physiology of skeletal muscle differs from that of liver in three ways important to our discussion of metabolic regulation (Fig. 15-18): (1) muscle uses its stored glycogen only for its own needs; (2) as it goes from rest to vigorous contraction, muscle undergoes very large changes in its demand for ATP, which is provided by glycolysis; (3) muscle lacks the enzymatic machinery for gluconeogenesis. The regulation of carbohydrate metabolism in muscle reflects these differences from liver. First, myocytes lack receptors for glucagon, which are present on hepatocytes. Second, the muscle isozyme of pyruvate kinase is not phosphorylated by PKA, so glycolysis is not turned off when [cAMP] is high. In fact, cAMP increases the rate of glycolysis in muscle, probably by activating glycogen phosphorylase. When epinephrine is released into the blood in a fight-or-flight situation, PKA is activated by the rise in [cAMP] and phosphorylates and activates glycogen phosphorylase kinase. The resulting phosphorylation and activation of glycogen phosphorylase results in faster glycogen breakdown. Epinephrine is not released under low-stress conditions, but with each neuronal stimulation of muscle contraction, cytosolic $[{Ca}^{2 +}]$ $left-bracket Ca Superscript 2 plus Baseline right-bracket$ rises briefly and activates phosphorylase kinase through its calmodulin subunit.

A figure compares the regulation of carbohydrate metabolism in the liver and a muscle. — FIGURE 15-18 Difference in the regulation of carbohydrate metabolism in liver and muscle. In liver, either glucagon (indicating low blood glucose) or epinephrine (signaling the need to fight or flee) has the effect of maximizing the output of glucose into the blood. In muscle, epinephrine increases glycogen breakdown and glycolysis, which together provide fuel to produce the ATP needed for muscle contraction.

Two vertical rectangles are shown with the left side labeled liver and the right side labeled muscle. Above the left side, the word glucagon has an arrow pointing downward. The word epinephrine above the center has an arrow pointing down to the liver side and a second arrow pointing down to the muscle side. In the liver side, glycogen is shown with an arrow down to glucose 6-phosphate. An arrow extends left out of the liver to a box labeled blood glucose. The same reaction is shown in the muscle side. Between them, the word glycogenolysis extends across both sides with an upward arrow on each side, one on the liver side and one in the muscle side. At the bottom of the liver side, the word pyruvate has an arrow pointing up to glucose 6-phosphate. To the right, the words glycolysis and gluconeogenesis are written in the center across both boxes. There is a downward arrow next to glycolysis and an upward arrow next to gluconeogenesis. At the bottom of the muscle side, an arrow points down from glucose 6-phosphate to pyruvate. To the left, there is an upward arrow next to glycolysis.

Elevated insulin triggers increased glycogen synthesis in myocytes by activating PP1 and inactivating GSK3. Unlike hepatocytes, myocytes have a reserve of the glucose transporter GLUT4 sequestered in intracellular vesicles. Insulin triggers their movement to the plasma membrane (see Fig. 12-23), where they allow increased glucose uptake. In response to insulin, therefore, myocytes help to lower blood glucose by increasing their rates of glucose uptake, glycogen synthesis, and glycolysis.

Carbohydrate and Lipid Metabolism Are Integrated by Hormonal and Allosteric Mechanisms

As complex as the regulation of carbohydrate metabolism is, it is far from the whole story of fuel metabolism. The metabolism of fats and fatty acids is very closely tied to that of carbohydrates. Hormonal signals such as insulin and changes in diet or exercise are equally important in regulating fat metabolism and integrating it with that of carbohydrates. We return to this overall metabolic integration in mammals in Chapter 23, after first considering the metabolic pathways for fats and amino acids (Chapters 17 and 18). The message we wish to convey here is that metabolic pathways are overlaid with complex regulatory controls that are exquisitely sensitive to changes in metabolic circumstances. These mechanisms act to adjust the flow of metabolites through various metabolic pathways, as needed by the cell and organism, without causing major changes in the concentrations of intermediates shared with other pathways.

SUMMARY 15.3 Coordinated Regulation of Glycogen Breakdown and Synthesis

Glycogen phosphorylase is activated in response to epinephrine (in muscle) or glucagon (in liver), which raise [cAMP] and thereby activate PKA. PKA phosphorylates and activates phosphorylase kinase, which converts glycogen phosphorylase b to its active a form. In muscle, ${Ca}^{2 +}$ $Ca Superscript 2 plus$ and AMP act allosterically to amplify the activity of the enzymes in this cascade. In liver, glucose acts allosterically to make phosphorylase a more susceptible to dephosphorylation/inactivation by phosphoprotein phosphatase 1 (PP1).
Glycogen synthase a is inactivated by phosphorylation, ultimately catalyzed by GSK3, but primed by phosphorylation by other kinases. Insulin, by causing inactivation of GSK3 and activating PP1, favors glycogen synthase a and stimulates glycogen synthesis. Glucose 6-phosphate acts allosterically to make glycogen synthase b a better substrate for PP1, which dephosphorylates it to its active a form.
In liver, glucagon stimulates glycogen breakdown and gluconeogenesis while blocking glycolysis, thereby sparing glucose for export to the brain and other tissues. In muscle, epinephrine stimulates glycogen breakdown and glycolysis, providing ATP to support contraction.
Further hormonal and allosteric mechanisms regulate the use of carbohydrates and lipids as metabolic fuels.