Berg J.M., Tymoczko J.L., Stryer L. Biochemistry

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21.4.2. Glycogen Synthase Catalyzes the Transfer of Glucose from UDP-Glucose to a

Growing Chain

New glucosyl units are added to the nonreducing terminal residues of glycogen. The activated glucosyl unit of UDP-

glucose is transferred to the hydroxyl group at a C-4 terminus of glycogen to form an α-1,4-glycosidic linkage. In

elongation, UDP is displaced by the terminal hydroxyl group of the growing glycogen molecule. This reaction is

catalyzed by glycogen synthase, the key regulatory enzyme in glycogen synthesis.

Glycogen synthase can add glucosyl residues only if the polysaccharide chain already contains more than four residues.

Thus, glycogen synthesis requires a primer. This priming function is carried out by glycogenin, a protein composed of

two identical 37-kd subunits, each bearing an oligosaccharide of α-1,4-glucose units. Carbon 1 of the first unit of this

chain, the reducing end, is covalently attached to the phenolic hydroxyl group of a specific tyrosine in each glycogenin

subunit. How is this chain formed? Each subunit of glycogenin catalyzes the addition of eight glucose units to its partner

in the glycogenin dimer. UDP-glucose is the donor in this autoglycosylation. At this point, glycogen synthase takes over

to extend the glycogen molecule.

21.4.3. A Branching Enzyme Forms α-1,6 Linkages

Glycogen synthase catalyzes only the synthesis of α-1,4 linkages. Another enzyme is required to form the α-1,6 linkages

that make glycogen a branched polymer. Branching occurs after a number of glucosyl residues are joined in α-1,4

linkage by glycogen synthase. A branch is created by the breaking of an α-1,4 link and the formation of an α-1,6 link:

this reaction is different from debranching. A block of residues, typically 7 in number, is transferred to a more interior

site. The branching enzyme that catalyzes this reaction is quite exacting. The block of 7 or so residues must include the

nonreducing terminus and come from a chain at least 11 residues long. In addition, the new branch point must be at least

4 residues away from a preexisting one.

Branching is important because it increases the solubility of glycogen. Furthermore, branching creates a large number of

terminal residues, the sites of action of glycogen phosphorylase and synthase (Figure 21.15). Thus, branching increases

the rate of glycogen synthesis and degradation.

Glycogen branching requires a single transferase activity. Glycogen debranching requires two enzyme activities: a

transferase and an α-1,6 glucosidase. Sequence analysis suggests that the two transferases and, perhaps, the α-1,6

glucosidase are members of the same enzyme family, termed the α -amylase family. Such an enzyme catalyzes a reaction

by forming a covalent intermediate attached to a conserved aspartate residue (Figure 21.16). Thus, the branching enzyme

appears to function through the transfer of a chain of glucose molecules from an α-1,4 linkage to an aspartate residue on

the enzyme and then from this site to a more interior location on the glycogen molecule to form an α-1,6 linkage.

21.4.4. Glycogen Synthase Is the Key Regulatory Enzyme in Glycogen Synthesis

The activity of glycogen synthase, like that of phosphorylase, is regulated by covalent modification. Glycogen synthase

is phosphorylated at multiple sites by protein kinase A and several other kinases. The resulting alteration of the charges

in the protein lead to its inactivation (Figure 21.17). Phosphorylation has opposite effects on the enzymatic activities of

glycogen synthase and phosphorylase. Phosphorylation converts the active a form of the synthase into a usually inactive

b form. The phosphorylated b form requires a high level of the allosteric activator glucose 6-phosphate for activity,

whereas the a form is active whether or not glucose 6-phosphate is present.

21.4.5. Glycogen Is an Efficient Storage Form of Glucose

What is the cost of converting glucose 6-phosphate into glycogen and back into glucose 6-phosphate? The pertinent

reactions have already been described, except for reaction 5, which is the regeneration of UTP. ATP phosphorylates

UDP in a reaction catalyzed by nucleoside diphosphokinase.

Thus, one ATP is hydrolyed incorporating glucose 6-phosphate into glycogen. The energy yield from the breakdown of

glycogen is highly efficient. About 90% of the residues are phosphorolytically cleaved to glucose 1-phosphate, which is

converted at no cost into glucose 6-phosphate. The other 10% are branch residues, which are hydrolytically cleaved. One

molecule of ATP is then used to phosphorylate each of these glucose molecules to glucose 6-phosphate. The complete

oxidation of glucose 6-phosphate yields about 31 molecules of ATP, and storage consumes slightly more than one

molecule of ATP per molecule of glucose 6-phosphate; so the overall efficiency of storage is nearly 97%.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.4. Glycogen Is Synthesized and Degraded by Different Pathways

Figure 21.15. Cross Section of a Glycogen Molecule. The component labeled G is glycogenin.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.4. Glycogen Is Synthesized and Degraded by Different Pathways

Figure 21.16. Structure of Glycogen Transferase.

A conserved aspartate residue forms a covalent intermediate with a

chain of glucose molecules.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.4. Glycogen Is Synthesized and Degraded by Different Pathways

Figure 21.17. Charge Distribution of Glycogen Synthase. Glycogen synthase has a highly asymmetric charge

distribution. Phosphorylation markedly changes the net charge of the amino- and carboxyl-terminal regions (yellow) of

the enzyme. The net charge of these regions and the interior of the enzyme before and after complete phosphorylation

are shown in green and red, respectively. [After M. F. Browner, K. Nakano, A. G. Bang, and R. J. Fletterick. Proc. Natl.

Acad. Sci. USA 86(1989):1443.]

II. Transducing and Storing Energy 21. Glycogen Metabolism

21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

We now return to the regulation of glycogen metabolism with a knowledge of both degradation and synthesis. Glycogen

breakdown and synthesis are reciprocally regulated by a hormone-triggered cAMP cascade acting through protein

kinase A (Figure 21.18). In addition to phosphorylating and activating phosphorylase kinase, protein kinase A adds a

phosphoryl group to glycogen synthase, which leads to a decrease in enzymatic activity. This important control

mechanism prevents glycogen from being synthesized at the same time that it is being broken down. How is the

enzymatic activity reversed so that glycogen breakdown halts and glycogen synthesis begins?

21.5.1. Protein Phosphatase 1 Reverses the Regulatory Effects of Kinases on Glycogen

Metabolism

The changes in enzymatic activity produced by protein kinases are reversed by protein phosphatases. The hydrolysis of

phosphorylated serine and threonine residues in proteins is catalyzed by protein phosphatases. One enzyme, termed

protein phosphatase 1, plays key roles in regulating glycogen metabolism. PP1 inactivates phosphorylase kinase and

phosphorylase a by dephosphorylating these enzymes. PP1 decreases the rate of glycogen breakdown: it reverses the

effects of the phosphorylation cascade. Moreover, PP1 also removes the phosphoryl group from glycogen synthase b to

convert it into the much more active a form. Hence, PP1 accelerates glycogen synthesis. PP1 is yet another molecular

device for coordinating carbohydrate storage.

The complete complex of PP1 consists of three components: PP1 itself, a 37-kd catalytic subunit; a 123-kd R

subunit

that confers high affinity for glycogen; and inhibitor 1, a small regulatory subunit that, when phosphorylated, inhibits

PP1. The importance of the R

subunit is that it brings PP1, which is active only when associated with glycogen

molecules, into proximity with its substrates.

How is the phosphatase activity of PP1 itself regulated? Consider the case in which glycogen degradation is predominant

(Figure 21.19). In this case, PKA is active. Two components of PP1 are themselves substrates for protein kinase A.

Phosphorylation of the R

component by protein kinase A prevents R

from binding the catalytic subunit of PP1.

Consequently, activation of the cAMP cascade leads to the inactivation of PP1 because it can no longer bind its

substrates. Phosphorylation of inhibitor 1 by protein kinase A blocks catalysis by PP1. Thus, when glycogen degradation

is switched on by cAMP, the accompanying phosphorylation of inhibitor 1 keeps phosphorylase in the active a form and

glycogen synthase in the inactive b form. The epinephrine-induced phosphorylation of the R

subunit and inhibitor 1 are

complementary devices for sustaining glycogen degradation.

21.5.2. Insulin Stimulates Glycogen Synthesis by Activating Protein Phosphatase 1

How is glycogen synthesis stimulated? As stated earlier, the presence of glucagon signifies the starved state and initiates

glycogen breakdown while inhibiting glycogen synthesis. When blood-glucose levels are high, insulin stimulates the

synthesis of glycogen by triggering a pathway that activates protein phosphatase 1 (Figure 21.20). The first step in the

action of insulin is its binding to a receptor tyrosine kinase in the plasma membrane. Multiple phosphorylations again

serve as the instigation for a regulatory wave of dephosphorylations. The binding of insulin to its receptor leads to the

activation of an insulin-sensitive protein kinase that phosphorylates the R

subunit of PP1 at a site different from that

modified by protein kinase A. This phosphorylation leads to the association of the R

subunit with PP1 and the

glycogen molecule. The consequent dephosphorylation of glycogen synthase, phosphorylase kinase, and phosphorylase

promotes glycogen synthesis and blocks its degradation. Once again we see that glycogen synthesis and breakdown are

coordinately controlled.

21.5.3. Glycogen Metabolism in the Liver Regulates the Blood-Glucose Level

After a meal rich in carbohydrates, blood-glucose levels rise, leading to an increase in glycogen synthesis in the liver.

Although insulin is the primary signal for glycogen synthesis, other, nonhormonal mechanisms also function in the liver.

One signal is the concentration of glucose in the blood, which normally ranges from about 80 to 120 mg per 100 ml (4.4

6.7 mM). The liver senses the concentration of glucose in the blood and takes up or releases glucose accordingly. The

amount of liver phosphorylase a decreases rapidly when glucose is infused (Figure 21.21). After a lag period, the amount

of glycogen synthase a increases, which results in the synthesis of glycogen. In fact, phosphorylase a is the glucose

sensor in liver cells. The binding of glucose to phosphorylase a shifts its allosteric equilibrium from the active R form to

the inactive T form. This conformational change renders the phosphoryl group on serine 14 a substrate for protein

phosphatase 1. It is significant that PP1 binds tightly to phosphorylase a but acts catalytically only when glucose induces

the transition to the T form. Recall that the R T transition of muscle phosphorylase a is unaffected by glucose and is

thus unaffected by the rise in blood-glucose levels (Section 21.2.2).

How does glucose activate glycogen synthase? Phosphorylase b, in contrast with phosphorylase a, does not bind the

phosphatase. Consequently, the conversion of a into b is accompanied by the release of PP1, which is then free to

activate glycogen synthase (Figure 21.22). Removal of the phosphoryl group of inactive glycogen synthase b converts it

into the active a form. Initially, there are about 10 phosphorylase a molecules per molecule of phosphatase. Hence, the

activity of glycogen synthase begins to increase only after most of phosphorylase a is converted into b. This remarkable

glucose-sensing system depends on three key elements: (1) communication between the serine phosphate and the

allosteric site for glucose, (2) the use of PP1 to inactivate phosphorylase and activate glycogen synthase, and (3) the

binding of the phosphatase to phosphorylase a to prevent the premature activation of glycogen synthase.

21.5.4. A Biochemical Understanding of Glycogen-Storage Diseases Is Possible

Edgar von Gierke described the first glycogen-storage disease in 1929. A patient with this disease has a huge

abdomen caused by a massive enlargement of the liver. There is a pronounced hypoglycemia between meals.

Furthermore, the blood-glucose level does not rise on administration of epinephrine and glucagon. An infant with this

glycogen-storage disease may have convulsions because of the low blood-glucose level.

The enzymatic defect in von Gierke disease was elucidated in 1952 by Carl and Gerty Cori. They found that glucose 6-

phosphatase is missing from the liver of a patient with this disease. This was the first demonstration of an inherited

deficiency of a liver enzyme. The liver glycogen is normal in structure but present in abnormally large amounts. The

absence of glucose 6-phosphatase in the liver causes hypoglycemia because glucose cannot be formed from glucose 6-

phosphate. This phosphorylated sugar does not leave the liver, because it cannot cross the plasma membrane. The

presence of excess glucose 6-phosphate triggers an increase in glycolysis in the liver, leading to a high level of lactate

and pyruvate in the blood. Patients who have von Gierke disease also have an increased dependence on fat metabolism.

This disease can also be produced by a mutation in the gene that encodes the glucose 6-phosphate transporter. Recall

that glucose 6-phosphate must be transported into the lumen of the endoplasmic reticulum to be hydrolyzed by

phosphatase (Section 16.3.5). Mutations in the other three essential proteins of this system can likewise lead to von

Gierke disease.

Seven other glycogen-storage diseases have been characterized (Table 21.1). In Pompe disease (type II), lysosomes

become engorged with glycogen because they lack α-1,4-glucosidase, a hydrolytic enzyme confined to these organelles

(Figure 21.23). The Coris elucidated the biochemical defect in another glycogen-storage disease (type III), which cannot

be distinguished from von Gierke disease (type I) by physical examination alone. In type III disease, the structure of liver

and muscle glycogen is abnormal and the amount is markedly increased. Most striking, the outer branches of the

glycogen are very short. Patients having this type lack the debranching enzyme (α-1,6-glucosidase), and so only the

outermost branches of glycogen can be effectively utilized. Thus, only a small fraction of this abnormal glycogen is

functionally active as an accessible store of glucose.

A defect in glycogen metabolism confined to muscle is found in McArdle disease (type V). Muscle phosphorylase

activity is absent, and the patient's capacity to perform strenuous exercise is limited because of painful muscle cramps.

The patient is otherwise normal and well developed. Thus, effective utilization of muscle glycogen is not essential for

life. The results of phosphorus-31 nuclear magnetic resonance studies of these patients have been very informative. The

pH of skeletal muscle cells of normal people drops during strenuous exercise because of the production of lactate. In

contrast, the muscle cells of patients with McArdle disease become more alkaline during exercise because of the

breakdown of creatine phosphate (Section 14.1.5). Lactate does not accumulate in these patients because the glycolytic

rate of their muscle is much lower than normal; their glycogen cannot be mobilized. The results of NMR studies have

also shown that the painful cramps in this disease are correlated with high levels of ADP (Figure 21.24). NMR

spectroscopy is a valuable, noninvasive technique for assessing dietary and exercise therapy for this disease.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.18. Coordinate Control of Glycogen Metabolism. Glycogen metabolism is regulated, in part, by hormone-

triggered cyclic AMP cascades: (A) glycogen degradation; (B) glycogen synthesis. Inactive forms are shown in red, and

active ones in green. The sequence of reactions leading to the activation of protein kinase A is the same in the regulation

of glycogen degradation and synthesis. Phosphorylase kinase also inactivates glycogen synthase.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.19. Regulation of Protein Phosphatase 1 (PP1). Phosphorylation of R

by protein kinase A dissociates the

catalytic subunit from the glycogen particle and hence the PP1 substrates. Inhibition is complete when the inhibitor

subunit (I) is phosphorylated and binds to PP1 to inactivate it.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.20. Insulin Activates Protein Phosphatase 1. Insulin triggers a cascade leading to the activation of protein

phosphatase 1, which results in the stimulation of glycogen synthesis and inhibition of its breakdown. The activated

receptor tyrosine kinase switches on a putative master kinase that phosphorylates the insulin-sensitive protein kinase. In

turn, the glycogen-targeting subunit (R

subunit) of the phosphatase is phosphorylated, which activates the enzyme.

[After P. Dent, A. Lavoinne, S. Nakielny, F. B. Caudwell, P. Watt, and P. Cohen. Nature 348(1990):306.]

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.21. Blood Glucose Regulates Liver Glycogen Metabolism. The infusion of glucose into the bloodstream

leads to the inactivation of phosphorylase, followed by the activation of glycogen synthase, in the liver. [After W.

Stalmans, H. De Wulf, L. Hue, and H.-G. Hers. Eur. J. Biochem. 41(1974):127.]

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.22. Glucose Regulation of Liver Glycogen Metabolism. Glucose binds to and inhibits glycogen

phosphorylase a in the liver, leading to the dissociation and activation of protein phosphatase 1 (PP1) from glycogen

phosphorylase a. The free PP1 dephosphorylates glycogen phosphorylase a and glycogen synthase b, leading to the

inactivation of glycogen breakdown and the activation of glycogen synthesis.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Table 21.1. Glycogen-storage diseases

Type Defective enzyme Organ

affected

Glycogen in the

affected organ

Clinical features

I Von Gierke

disease

Glucose 6-phosphatase or transport

system

Liver and

kidney

Increased amount;

normal structure.

Massive enlargement of the

liver. Failure to thrive.

Severe hypoglycemia,

ketosis, hyperuricemia,

hyperlipemia.

II Pompe

disease

α-1,4-Glucosidase (lysosomal)

All organs Massive increase

in amount; normal

structure.

Cardiorespiratory failure

causes death, usually before

age 2.

III Cori disease Amylo-1,6-glucosidase

(debranching enzyme)

Muscle and

liver

Increased amount;

short outer

branches.

Like type I, but milder

course.

IV Andersen

disease

Branching enzyme

Liver and

spleen

Normal amount;

very long outer

branches.

Progressive cirrhosis of the

liver. Liver failure causes

death, usually before age 2.

V McArdle

disease

Phosphorylase Muscle Moderately

increased amount;

normal structure.

Limited ability to perform

strenuous exercise because of

painful muscle cramps.

Otherwise patient is normal

and well developed.

VI Hers disease Phosphorylase Liver Increased amount. Like type I, but milder

course.

VII Phosphofructokinase Muscle Increased amount;

normal structure.

Like type V.

VIII Phosphorylase kinase Liver Increased amount;

normal structure.

Mild liver enlargement. Mild

hypoglycemia.

Note: Types I through VII are inherited as autosomal recessives. Type VIII is sex linked.

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.23. Glycogen-Engorged Lysosome. This electron micrograph shows skeletal muscle from an infant with type

II glycogen-storage disease (Pompe disease). The lysosomes are filled with glycogen because of a deficiency in α-1,4-

glucosidase, a hydrolytic enzyme confined to lysosomes. The amount of glycogen in the cytosol is normal. [From H.-G.

Hers and F. Van Hoof, Eds. Lysosomes and Storage Diseases (Academic Press, 1973), p. 205.]

II. Transducing and Storing Energy 21. Glycogen Metabolism 21.5. Glycogen Breakdown and Synthesis Are Reciprocally Regulated

Figure 21.24. NMR Study of Human Arm Muscle. The level of ADP during exercise increases much more in a patient

with McArdle glycogen-storage disease (type V) than in normal controls. [After G. K. Radda. Biochem. Soc. Trans. 14

(1986):522.]

II. Transducing and Storing Energy 21. Glycogen Metabolism

Summary

Glycogen, a readily mobilized fuel store, is a branched polymer of glucose residues. Most of the glucose units in

glycogen are linked by α-1,4 glycosidic bonds. At about every tenth residue, a branch is created by an α-1,6-glycosidic

bond. Glycogen is present in large amounts in muscle cells and in liver cells, where it is stored in the cytoplasm in the

form of hydrated granules.

Glycogen Breakdown Requires the Interplay of Several Enzymes

Most of the glycogen molecule is degraded to glucose 1-phosphate by the action of glycogen phosphorylase, the key

enzyme in glycogen breakdown. The glycosidic linkage between C-1 of a terminal residue and C-4 of the adjacent one is

split by orthophosphate to give glucose 1-phosphate, which can be reversibly converted into glucose 6-phosphate.

Branch points are degraded by the concerted action of an oligosaccharide transferase and an α-1,6-glucosidase.

Phosphorylase Is Regulated by Allosteric Interactions and Reversible Phosphorylation

Phosphorylase is regulated by allosteric effectors and reversible covalent modifications. Phosphorylase b, which is

usually inactive, is converted into active phosphorylase a by the phosphorylation of a single serine residue in each

subunit. This reaction is catalyzed by phosphorylase kinase. The b form in muscle can also be activated by the binding of

AMP, an effect antagonized by ATP and glucose 6-phosphate. The a form in the liver is inhibited by glucose. The AMP-

binding sites and phosphorylation sites are located at the subunit interface. In muscle, phosphorylase is activated to

generate glucose for use inside the cell as a fuel for contractile activity. In contrast, liver phosphorylase is activated to

liberate glucose for export to other organs, such as skeletal muscle and the brain.

Epinephrine and Glucagon Signal the Need for Glycogen Breakdown

Epinephrine and glucagon stimulate glycogen breakdown through specific 7TM receptors. Muscle is the primary target

of epinephrine, whereas the liver is responsive to glucagon. Both signal molecules initiate a kinase cascade that leads to

the activation of glycogen phosphorylase.

Glycogen Is Synthesized and Degraded by Different Pathways