Glycolysis, gluconeogenesis, glycogen metabolism, TCA cycle, and PPP

METABOLISM OF CHAPTER
C ARBOHYDRATES I:
MAINLINE METABOLIC
PATHWAYS 9
Carbohydrates are important for both the generation of metabolic energy and biosynthetic purposes. They are the struc-
tural basis of cellulose, bones, cartilage, lubricants such as mucus, cell recognition, DNA, RNA, and some membrane lipids
(sphingosine). They are source of carbon skeleton of certain amino acids and are basis of some intracellular messenger
systems. As an energy source they are broken down, and when there is a surplus, they are stored as starch and glycogen.
Glucose occupies a key position in carbohydrate metabolism, being a building block of all major dietary carbohy-
drates and the principal transported carbohydrate in humans. Fructose and galactose, obtained from dietary sucrose and
lactose respectively, can be converted to glucose in liver. Therefore, the largest portion of dietary carbohydrates reaches
the consuming tissues in the form of glucose. Evidently, an understanding of glucose metabolism forms the core of study
of carbohydrate metabolism.
A detailed account of metabolic pathways is given in this chapter. After going through this chapter, the student should
be able to understand:
Transport of glucose into cells: facilitated transport and secondary active transport across cell membrane.
Pathway of glycolysis: reaction sequence, generation of ATP and regulation.
Feeder pathways: reaction sequences for monosaccharides, disaccharides and polysaccharides, and the associated
metabolic diseases.
Tricarboxylic acid cycle: reactions, energy yield, synthetic function and regulation.
Pathway of gluconeogenesis; bypass reactions and reversible steps; energetics and regulation.
Glycogen metabolism reactions of glycogen synthesis (glycogenesis) and degradation (glycogenolysis), their reciprocal
regulation through cAMP activated cascade; role of hormones in the regulation; and biochemical basis and clinical
presentations of glycogen storage diseases.
I. Transport of Glucose Into Cells Facilitated transport occurs along a concentration gra-
dient and is mediated via a family of at least five transport
Glucose cannot passively move across the cell membrane proteins located in the cell membrane. These proteins are
to enter the cells. The polar nature of glucose hinders its designated as GLUT-1, GLUT-2, GLUT-3 GLUT-4 and
movement across the predominantly non-polar lipid GLUT-5. Glucose moves from higher extracellular glucose
bilayer. Two kinds of transport mechanisms are present in concentration to a lower concentration inside the cell. A
the cell membrane, which permit glucose to cross the mem- given transport protein exists in two conformational states,
branous barrier and enter the cell. These are (a) the facil- which alternate during the transport process (Fig. 7.11).
itated transport, and (b) the secondary active transport. The glucose transport proteins are tissue specific.
Both are carrier-mediated processes, requiring transport GLUT-1 is present in RBCs, whereas GLUT-4 is abundant
protein(s). in skeletal muscles and adipose tissue. Insulin increases
158 Textbook of Medical Biochemistry
number and activity of GLUT-4, thereby promoting entry Uronic acid pathway
of glucose in these tissues (Chapter 7).
Note: Insulin is not required for glucose uptake by some
Glucose Glucose 6-phosphate Glycolysis
tissues, such as liver, brain and red blood cells.
Secondary active transport is the mechanism of sodium-
glucose cotransport across the membranous barrier, against Liver
Pi HMP shunt
a concentration gradient of glucose (i.e. from lower glucose Glucogenesis
concentration to a relatively higher concentration). The Glycogenolysis
“uphill movement” of glucose is powered by the movement
Fig. 9.1. The role of glucose 6-phosphate in carbohydrate
of sodium along its concentration gradient (Chapter 7).
metabolism.
This process is operative during the movement of glucose
from lumen of intestine into the intestinal mucosal cells Glucose
(Fig. 7.16). Another type of such Na/glucose co-transporter
Glycolysis
(called SLUT-1) is known to operate in renal tubules.
Abnormalities in some of the transport proteins may
Pyruvate
lead to diseases; for example, renal glycosuria results in case
An
of a defect in the renal transport mechanism for glucose. bic co aero
Mitochondrial e ro ions nd bi
A dit itio c
matrix
co
n ns
Pyruvate
Specific integral membrane proteins facilitate movement
of glucose along concentration gradient, without expen- Lactate
diture of any energy in facilitated transport; whereas in
secondary active transport a specific transport protein Acetyl CoA
moves glucose against concentration gradient through
expenditure of energy; the energy comes by co-transport
of sodium ions along its concentration gradient. TCA
Inner Cytosol
cycle
mitochondrial
Glucose 6-phosphate: a branch-point compound: After membrane
entering the cell glucose is rapidly converted to glucose CO2 + H2O
6-phosphate. The latter serves as a common link between
Fig. 9.2. Pyruvate metabolism under aerobic and anaerobic
various metabolic pathways, therefore called a branch conditions. Under anaerobic conditions, pyruvate is reduced to
point compound. Figure 9.1 shows some important inter- lactate, whereas in aerobic conditions pyruvate is oxidized to
connections. Glucose 6-phosphate can enter a number of acetyl CoA, which is further oxidized in TCA cycle.
pathways, e.g. glycolysis, uronic acid pathway, HMP shunt,
glycogenesis, and can again form glucose by hydrolytic important. This pathway is known as glycolysis or the
removal of the phosphate group. It can be generated by Embden-Meyerhof pathway. It is an anaerobic process that
glycogenolysis and gluconeogenesis also. Further, glucose involves the breaking down of a molecule of glucose into two
6-phosphate cannot simply diffuse out of the cell by cross- molecules of pyruvate or lactate.
ing the cell membrane and hence is committed to intracel- Glucose (2) Pyruvate/Lactate
luar metabolism. Detailed description of various pathways (6-c) (3-c) (3-c)
mentioned above has been given in Chapters 9 and 10. Glycolysis is common to most organisms, and in
humans it occurs virtually in all tissues. Although the gly-
Glucose enters the call by a carrier-mediated transport colytic sequence usually begins with glucose, other sugars
mechanism (facilitated transport or secondary active may also enter it via appropriate intermediates. Glycolysis
transport), where it is rapidly phosphorylated to glu- serves a dual role:
cose 6-phosphate—a compound that can enter several to generate metabolic energy
metabolic pathways. to provide intermediates for several other metabolic
pathways.
Glycolysis occurs in 10 sequential reactions, that occur
II. Glycolysis in the cytosol. The fate of pyruvate, the end product of
these reactions, is different in anaerobic and aerobic
Of all the metabolic transformations that glucose 6- conditions (Fig. 9.2).
phosphate can undergo, the sequence of reactions leading Under aerobic conditions: Pyruvate enters the mito-
to the formation of pyruvate is quantitatively the most chondrial matrix and is oxidized to acetyl CoA, which is the
Metabolism of Carbohydrates I: Mainline Metabolic Pathways 159
major metabolic fuel for the citric acid cycle. Further oxida- Glucokinase, on the other hand, is specific for glucose.
tion of acetyl CoA in the citric acid cycle releases free energy It has high Vmax and, unlike hexokinase, is not inhibited by
which is used to generate ATP molecules. This process the reaction product, glucose 6-phosphate. Moreover, it
occurs in all tissues except those which lack mitochondria has high Km (20 mmol/L), i.e. low affinity for glucose and
(e.g. erythrocytes, leucocytes) and the exercising muscles. therefore, comes into play only when intracellular glu-
Under anaerobic conditions: Pyruvate is reduced in cose concentration rises steeply, such as following a car-
cytosol by NADH to form lactate. This process, called bohydrate rich diet. These properties make the glucokinase
anaerobic glycolysis is discussed later. (Note that the term suitable for rapidly phosphorylating the glucose that is
anaerobic literally means without air, but in practice it presented to liver following a meal. Within the hepato-
means without oxygen). cytes, the glucose molecules are rapidly phosphorylated by
glucokinase (due to high Vmax and high Km), without any
hindrance to its action (due to lack of feedback inhibition).
Glycolysis is a cytoplasmic reaction sequence of 10 The above transformation is further enhanced because of
enzyme-catalyzed reactions which produces two mole- induction of glucokinase synthesis after a meal, especially
cules of the three-carbon compound, pyruvate (or lac-
following a carbohydrate rich diet. The enzyme induction
tate), from the six-carbon substrate, glucose.
is probably brought about by insulin, that is released at
such time. Large quantities of glucose 6-phosphate are
thereby produced, which are channeled into the pathway
A. Reactions of glycogen synthesis (i.e. glycogenesis).
Glycolysis can be divided into two main stages. The stage

I, which comprises the first five reactions, is an endergonic Glucokinase can rapidly phosphorylate large amount
(i.e. energy-requiring) stage in which glucose molecules of glucose after meals, which then enters the pathway
are activated through the expenditure of ATP molecules. of glycogen synthesis.
Thus, this stage involves energy investment. Stage II leads
to energy generation and hence referred to as the “payoff Reaction 2: Isomerization of Glucose 6-Phosphate
stage“. Glucose 6-phosphate, an aldose sugar, is now prepared
Reactions of glycolysis are shown in Figure 9.3. for subsequent cleavage into two 3-carbon molecules.
To split anything is easier if it is bilaterally symmetrical.
Stage I: The Investment Phase Glucose 6-phosphate is, however, not very symmetrical,
Reaction 1: Phosphorylation of Glucose and so converted to fructose 6-phosphate, a little more
Glucose is phosphorylated at the sixth carbon to form symmetrical molecule. It is a freely reversible reaction
glucose 6-phosphate. This places a large negative charge catalyzed by phosphohexose isomerase.
on the glucose molecule so that it cannot simply diffuse Reaction 3: Phosphorylation of Fructose 6-Phosphate
out of the cell. The phosphate group is obtained from Fructose 6-phosphate, formed in the previous step, is
ATP which is converted to ADP in the process, which made more symmetrical by esterifying a phosphate to
means that energy is lost in this step. However, this is C-1. This yields fructose 1,6-bisphosphate, a molecule
better than losing glucose out of the cell. that is almost symmetrical. The phosphate group comes
from ATP and the enzyme is phosphofructokinase (PFK1).
Glucose ATP Glucose 6-phosphate ADP
Note: A diphosphate differs from bisphosphate (and a tri-
The enzyme involved in this reaction is kinase, (an
phosphate from trisphosphate) in that, for a diphosphate
enzyme that catalyzes a phosphorylation and uses ATP as
(e.g. ADP) the phosphates are joined to each other, whereas
its source of phosphate is called kinase). There are two
in a bisphosphate (e.g. fructose 1, 6-bisphosphate), the
possible enzymes for this reaction, depending on the
phosphate groups are attached at different places on the
tissue. One is glucokinase (found in liver) and the other is
sugar.
hexokinase (muscle and fat), which will catalyze phosphor-
ylation of most hexoses, including glucose. Kinetic prop- This reaction is irreversible and is the “rate-limiting step“
erties of the two enzymes are different (Table 9.1). for glycolysis. It is also the committed step, meaning that
Hexokinase has low Km, i.e. high affinity for glucose, low once fructose 1, 6-bisphosphate is formed it must go for
Vmax, and is subject to feedback inhibition by the reaction the glycolytic pathway only. Activity of PFK1 is controlled
product, glucose 6-phosphate. The hexokinase reaction is by a variety of positive and negative regulatory modula-
not specific for glycolysis; it only commits glucose to tors, which make this step the most important regulatory
intracellular metabolism because glucose 6-phosphate is step of the pathway. Regulation of glycolysis is discussed
not transported across the plasma membrane. later in this chapter.
H O H O
CH2OH
C Hexokinase C
C O
(glucokinase in liver) Phosphohexose
H C OH H C OH
isomerase HO C H
HO C H HO C H
1 H C OH
ATP ADP H C OH 2
H C OH H C OH
H C OH H C OH 2−
CH2OPO3
2−
CH2OH CH2OPO3
Fructose 6-phosphate
D-Glucose Glucose 6-phosphate
3 2−
CH2OPO3
ATP
Phosphofructokinase C O
CH2OH
Dihydroxyacetone
ADP 4 Aldolase phosphate Triose
2− phosphate
CH2OPO3 H O
isomerase
C O C
HO C H H C OH 5
2−
H C OH CH2OPO3
H C OH (2x) Glyceraldehyde 3-phosphate
2− Pi
CH2OPO3
Fructose NAD+ Glyceraldehyde
6 3-phosphate
1,6-Bisphosphate
dehydrogenase
NADH+H+
O
2−
C OPO3
H C OH
2−
CH2OPO3
(2x) 1,3-Bisphosphoglycerate
ADP
Phosphoglycerate
7 kinase
ATP
O
C O−
H C OH
2−
CH2OPO3
9 (2x) 3-phosphoglycerate
10
O H2O 8 Phosphoglycerate
ATP ADP mutase
O C O– O
(2x) Acetyl CoA
(2x) Pyruvate C O– C
2−
OPO3 C O−
(2x) Lactate Pyruvate Enolase 2−
(2x) Alanine C O CH2 H CH2OPO3
kinase
(2x) Oxaloacetate CH3 (2x) Phosphoenolpyruvate CH2OH
(2x) Ethanol
(2x) 2-Phoshoglycerate
Fig. 9.3. The pathway of glycolysis and fate of pyruvate. Stage I: Reactions 1–5—energy-consuming reactions and Stage II:
6–10—energy-yielding reactions. Irreversible steps: 1st, 3rd and 10th).
Table 9.1. Comparative properties of hexokinase and Reaction 4: Cleavage of Fructose 1,6-Bisphosphate
glucokinase Fructose 1,6-bisphosphate, a 6-carbon compound, is
cleaved by the enzyme aldolase into two trioses: glyceral-
Hexokinase Glucokinase
dehyde 3-phosphate and dihydroxyacetone phosphate.
Specificity Broad (hexoses) Narrow (glucose only)
This enzyme, which splits an aldose, is not specific for
Feedback inhibition Yes No
fructose 1,6-bisphosphate, hence its general name.
Affinity for glucose High Low
Km value 10–2 mmol/L 20 mmol/L Reaction 5: Isomerization of Dihydroxyacetone Phosphate
Of the two trioses, only glyceraldehyde 3-phosphate can
Vmax value Low High
be further metabolized through the glycolytic sequence.
Dihydroxyacetone phosphate, therefore, must be con- by transferring the phosphate to ADP to form ATP (i.e.
verted to glyceraldehyde 3-phosphate for further metab- substrate level phosphorylation).
olism. This aldose-ketose isomerization is catalyzed by This reaction is catalyzed by the enzyme phosphoglycer-
the enzyme, triose phosphate isomerase. ate kinase. Since two molecules of 1,3-bisphosphoglycer-
Fructose 1,6-bis p Dihydroxyacetone- p ate are produced from a single molecule of glucose, two
+ ATPs are actually generated in this step from a single glu-
Glyceraldehyde 3- p
cose molecule.
Sum: Fructose 1,6-bis- p (2x) Glyceraldehyde 3- p The 1,3-BPG is converted to an alternate metabolite,
It is necessary to understand that two glyceraldehyde 2,3-bisphosphoglycerate (2,3-BPG) in erythrocytes which
3-phosphate molecules are generated from the original enhances unloading of oxygen (Box 9.1).
glucose molecule and enter this part of the pathway, and
therefore, all of the following steps occur twice, once for each
Reaction 8: Inter-Molecular Shift of Phosphate Group
The phosphate group is transferred from the C-3 of
of the two glyceraldehyde 3-phosphate molecules.
the 3-phosphoglycerate to the C-2 by the enzyme
phosphoglycerate mutase, resulting in the formation of
In the first stage of glycolysis, glucose is phosphorylated, 2-phosphoglycerate.
isomerized, phosphorylated again and cleaved to yield Note: The term mutase is used to designate the enzyme
two triose molecules. These reactions consume 2ATPs per
catalyzing transpositioning of functional group, within
glucose. All reactions beyond this point occur twice for
the same molecule.
each glucose molecule.
Reaction 9: Dehydration of 2-Phosphoglycerate
Removal of a water molecule from 2-phosphoglycerate is
Stage II: Energy Pay-off Phase catalyzed by the enzyme enolase, resulting in formation
The next few steps are remarkable for they bring about of phosphoenolpyruvate (PEP). In this reaction, redistri-
oxidation of each of the three carbons of glyceralde- bution of energy occurs within the same molecule, so that
hydes. There is addition of oxygen at C-1, loss of hydro- the phosphate ester (PEP) becomes unusually energy rich
gen at C-2, and loss of phosphate at C-3. In these steps a (G0‘ 14.8 kcal/mole), which is sufficient for genera-
large amount of energy is liberated, enough to replace tion of ATP (i.e. substrate level phosphorylation).
the energy lost so far (2ATPs) plus some more. Inhibition of enolase catalyzing this step has impor-
Reaction 6: Dehydrogenation of Glyceraldehyde tant role in clinical biochemistry (Box 9.2).
3-Phosphate Reaction 10: ATP Production from Phosphoenol Pyruvate
Dehydrogenation of glyceraldehyde 3-phosphate is cata- The last step of the glycolytic sequence involves transfer
lyzed by the enzyme glyceraldehyde 3-phosphate dehydrogenase, of high energy phosphate group from PEP to ADP to
resulting in the formation of 1,3-bisphosphoglycerate. yield a molecule of ATP (Fig. 9.4).
Pi, NAD+ NADH, H+
This is the third irreversible step of glycolysis (reaction
1 and 3 are the other two). Pyruvate kinase, the enzyme
Glyceraldehyde 3- 1,3-Bisphospho- catalyzing this step, is a regulatory enzyme.
phosphate glycerate Deficiency of pyruvate kinase causes decreased
production of ATP from glycolysis. Red blood cells are
There are two interesting aspects of this reaction: predominantly affected in this disorder. They are unable
firstly, the pair of hydrogen atoms removed is replaced to generate sufficient ATP for their sodium pumps.
by a phosphate group. This hydrogen is transferred to Consequently, the ionic gradient across the erythrocyte
NAD to form NADH, which is a valuable product (its membrane cannot be maintained, resulting in lysis of
oxidation in the respiratory chain is a source of ATP). the cell. This results in haemolysis, and hence haemolytic
Second, the phosphate group added to the glyceralde- anaemia.
hyde 3-phosphate molecules is inorganic phosphate.
This means that energy is not lost, which would have
ADP ATP
been the case if it had come from ATP. COO− O− COO− COO−
Reaction 7: ATP Production from 1,3-Bisphosphoglycerate C O P O− C OH C O

The 1,3-bisphosphoglycerate (1,3-BPG) contains a carbox- CH2 O CH2 CH3
Phosphoenolpyruvate (PEP) Enolpyruvate Pyruvate
ylic anhydride with phosphate at C-1. This type of anhy-
dride, called acyl-phosphate, has a very high standard free Fig. 9.4. The pyruvate kinase reaction. Pyruvate shows keto-
energy of hydrolysis; the G0’ of 1,3-bisphosphoglycerate enol tautomerism, the keto form being energetically far more
is 11.8 kcal/mole (Chapter 8). The cell uses this energy stable than the enol form.
BOX 9.1
Role of 2,3-Bisphosphoglycerate in Erythrocytes
In erythrocytes, 1,3-BPG is converted to 2,3-bisphosphoglycerate (2,3-BPG) by catalytic action of the enzyme 2,3-bisphos-
phoglycerate mutase. The 2,3-BPG concentration sometimes reaches 5 mmol/L, comparable with the molar concentration
of haemoglobin in the RBC, and much higher than that of ATP (1–2 mmol/L) or inorganic phosphate (1 mmol/L). 2,3-BPG is
a negative allosteric effector of the oxygen affinity of haemoglobin: it decreases the affinity to promote the release of
oxygen (from oxyhaemoglobin) in peripheral tissues. Its concentration increases in the RBCs, during adaptation to higher
altitudes and in anaemia, thus promoting the release of oxygen to tissues when the oxygen transport capacity of blood is
decreased. Fetal haemoglobin is less sensitive than adult haemoglobin to the effects of this compound, so that 2,3-BPG in mater-
nal erythrocytes is one factor that promotes efficient transfer of oxygen across the placenta from HbA to HbF (Chapter 17).
Finally, 2,3-BPG is hydrolyzed to 3-phosphoglycerate by bisphosphoglycerate phosphatase. It is postulated that it is a
bifunctional enzyme catalyzing both the synthesis of 2,3-BPG (from 1,3-BPG) and its subsequent hydrolysis.
More detailed discussion on 2,3 BPG-is given in Chapter 17.
Note: This pathway of synthesis and breakdown of 2,3-BPG in erythrocytes causes bypassing of the glycerophosphate
kinase reaction (step 6 of glycolysis) and is referred to as Rapaport–Leubering cycle (or simply BPG cycle). In this shunt
pathway, no ATP is generated. Normally, about 15–25% of the glucose converted to lactate in erythrocytes is routed via
this cycle. This is useful not only in the oxygen unloading by oxyhaemoglobin, but also in dissipation of energy, and there-
fore, advantageous to the cell when the energy requirement is minimal.
BOX 9.2
Inhibition of Glycolysis
The glycolytic enzyme enolase is inhibited by fluoride, which thereby inhibits the glycolytic sequence. Sodium fluoride is
mixed with potassium oxalate, an anticoagulant, in the blood sample collected for glucose estimation. Because it inhibits
enolase, in vitro glycolysis is prevented. In absence of sodium fluoride, glucose will be used.
Some other inhibitors of glycolysis are as below:
1. Iodoacetate, which inhibits the enzyme glyceraldehyde 3-phosphate dehydrogenase.
2. Arsenite, which inhibits phosphoglycerate kinase.
2. Reduction to lactate: Lactate is produced by glycolysis

In the second stage of glycolysis, a series of changes under anaerobic conditions and the process is called
convert glyceraldehyde 3-phosphate to pyruvate. This anaerobic glycolysis. The pyruvate to lactate reduc-
stage produces 4 ATPs per glucose molecule for a net tion is brought about by the cytosolic enzyme lactate
yield of 2 ATPs per glucose. dehydrogenase and requires NADH.
NADH, H+ NAD+
B. Fate of Pyruvate Pyruvate Lactate

Lactate
dehydrogenase
There are several pathways into which pyruvate can enter
(Fig. 9.3). The pathway chosen in a given tissue depends The NADH is produced in the glyceraldehyde 3-phosphate
on its state of oxygenation and the prevailing metabolic dehydrogenase reaction (Reaction 6) of glycolysis (Fig. 9.5).
conditions, as described below: The pyruvate to lactate conversion regenerates NAD for
1. Oxidative decarboxylation to acetyl CoA: In tissues that the above reaction so that glycolysis can proceed nor-
are adequately perfused with oxygen (i.e. under aero- mally. It is noteworthy, that without regeneration of
bic conditions), pyruvate undergoes oxidative decar- NAD by lactate dehydrogenase reaction, the Reaction 6
boxylation to form acetyl CoA, which is further would stop and the glycolytic pathway would also soon
oxidized via citric acid cycle. be halted for lack of NAD. Thus anaerobic glycolysis
Glucose Acetyl CoA

Serine
−2ATP Lactate
Thronine
Alanine Oxaloacetate
Pyruvate Malate
Malate
+4ATP Alanine
Glucose
Ethanol
Lactate
COO− NAD+ NADH, H+ COO−
Fig. 9.6. Pyruvate can be generated from several compounds,
HC OH C O
Lactate and can be channeled into various pathways.
CH3 dehydrogenase CH3
Lactate Pyruvate
Finally, during hypoxia resulting from vascular obstruc-
Fig. 9.5. Outline of reactions of anaerobic glycolysis. It regen- tion or other causes, for example, in acute myocardial
erates NAD (for the glyceraldehyde 3-phosphate dehydrogenase infarction – anaerobic glycolysis occurs to help the cells
reaction) so that glycolytic sequence can proceed smoothly survive the brief episodes of oxygen depletion.
with generation of 2ATP molecules.
Significance
provides elegantly simple solution to ensure that glycoly- Anaerobic glycolysis provides less energy: net gain of
sis proceeds uninterruptedly. ATPs is only two molecules (Fig. 9.5). However, it is an
More about anaerobic glycolysis is discussed at end of important process because of two reasons:
this section.
1. NAD is generated during their process, which is
required for step 6 of glycolysis (Fig. 9.5). As discussed
Under aerobic conditions, pyruvate enters the mitochon- in the previous section, NAD regeneration must occur
drion, where it is oxidatively decarboxylated to acetyl
for, glycolysis to continue.
CoA, and under anaerobic conditions, pyruvate is
2. Carbohydrates are the only metabolic substrates that
reduced to regenerate NAD+ for glycolysis.
can produce ATP under anaerobic conditions.
3. Carboxylation to oxaloacetate: Pyruvate is converted to
oxaloacetate by biotin-dependent carboxylation cata- C. Generation of ATP by Glycolysis
lyzed by the enzyme pyruvate carboxylase. The oxaloac-
etate can form glucose via gluconeogenesis. Energy yield in the anaerobic and the aerobic states are
4. Conversion to malate: Malic enzyme converts pyruvate different, and therefore, discussed separately.
to malate, which can also enter gluconeogenic
sequence.
Aerobic State
5. Conversion to alanine: Pyruvate may form the amino
In course of conversion of one molecule of glucose
acid, alanine by transamination.
through glycolytic sequence to two molecules of pyru-
6. Reduction to ethanol: Conversion of pyruvate to ethanol
vate, there occurs generation of four molecules of ATP
occurs in yeasts and some bacteria, including those of
and, utilization of two molecules of ATP.
the intestinal flora.
Initially, two ATPs are utilized to phosphorylate glu-
Pyruvate can be funneled into various metabolic path- cose to glucose 6-phosphate and fructose 6-phosphate to
ways. It can also be generated from a number of com- fructose 1,6-bisphosphate (see reactions 1 and 3; Fig. 9.3).
pounds, as shown in Figure 9.6. Evidently, pyruvate is said Subsequently, cleavage of fructose 1,6-bisphosphate, fol-
to be at the metabolic cross-roads. lowed by isomerization of dihydroxyacetone phosphate,
produces two molecules of glyceraldehyde 3-phosphate.
Anaerobic Glycolysis Therefore, from each molecule of glucose, two molecules
Anaerobic glycolysis occurs in tissues that lack mito- of triose phosphate are produced. Each molecule of tri-
chondria, such as erythrocytes, lens and cornea of the eye, ose phosphate then generates two molecules of ATP: one
renal medulla, and leukocytes. In oxygen deficient tissues ATP is produced by phosphoglycerate kinase (Reaction 7)
such as severely exercising muscles, this pathway is cho- and the other by pyruvate kinase (Reaction 10). Therefore,
sen. Oxygen deficiency occurs because the blood is two molecules of triose-phosphate generates four ATPs.
squeezed out of the contracting muscles. Moreover, pro- With two molecules of ATP invested and four generated, net
duction of pyruvate in exercising muscle is very fast due gain of ATPs is two molecules (Table 9.2).
to rapid rate of glycolysis, but its oxidation is much In addition to these, generation of more ATPs occurs by
slower due to oxygen deficiency. The surplus pyruvate is oxidation of the NADH, (generated earlier in glyceralde-
diverted for lactate formation (Fig. 9.5). hyde 3-P dehydrogenase reaction). Each of these NADH
Table 9.2. Energy yield during the conversion of one molecule LIVER MUSCLE
of glucose to two molecules of pyruvate in aerobic glycolysis*
Glucose Glucose
Reaction Enzyme Product
1st Hexokinase 1 ATP Gluconeo- Glycolysis
genesis
3rd Phosphofructokinase 1 ATP
6th Glyceraldehyde 3-phosphate 2 NADH BLOOD
Pyruvate Pyruvate
dehydrogenase NADH + H+ Lactate NADH + H+
Lactate
7th Phosphoglycerate kinase 2 ATP dehydrogenase dehydrogenase
NAD+ NAD+
10th Pyruvate kinase 2 ATP
2 ATP 2 NADH Lactate Lactate
* Note that all reactions beyond the aldolase reaction occur twice for
each glucose molecule. Fig. 9.7. Cori cycle. The exercising muscles generate lactate
via anaerobic glycolysis, which is transported to liver for
resynthesis of glucose.
molecules yields three ATPs by oxidative phosphorylation
(Chapter 14). Thus, a total of six ATPs are produced from Cori and Gerty Cori who were awarded Nobel prize in
this source. When two ATPs generated earlier are added, 1947 for this discovery.
the net production of ATPs in aerobic glycolysis becomes eight.

Vigorously exercising muscles generate lactate via anaerobic
Anaerobic State glycolysis. During recovery, some of this lactate is trans-
Generation of two molecules of ATP and two molecules of ported to liver and used to form glucose via gluconeogen-
NADH occurs initially in this type also. The latter are oxi- esis. The overall pathway (glucose lactate glucose)
dized to NAD during reduction of pyruvate. Thus, there is constitutes the Cori cycle.
no net production of NADH (Fig. 9.5) in anaerobic glycoly-
sis and, therefore, net generation of only two ATP occurs.
Lactate, the end product of anaerobic glycolysis, still D. Regulation
contains a large amount of inherent free energy, but is
not degraded further. Thus anaerobic glycolysis releases Glycolysis is central to metabolism and is integrated with
only a small fraction of the energy of the glucose mole- a number of other metabolic pathways. Therefore, the
cules. Still it is a valuable source of energy in tissues lack- regulatory enzymes of glycolysis respond to appropriate
ing mitochondria. Moreover, in case of oxygen depletion, signals from several other pathways. This makes the regu-
the affected tissues depend more on anaerobic glycolysis lation of glycolysis a complex event. There are three
for getting the required energy. Production of lactate is glycolytic reactions catalyzed by hexokinase (or glucoki-
thereby increased. However, excessive production of lac- nase), phosphofructokinase (PFK1) and pyruvate kinase,
tate may have serious consequences (Case 9.1). which are metabolically irreversible and subject to regu-
lation. The main control step is that catalyzed by PFK1 but
Cori Cycle or Lactic Acid Cycle hexokinase and pyruvate kinase are additional control sites.
In an actively exercising muscle, less than 10% of pyru-
vate is utilized by the citric acid cycle, and the rest is Regulation of Hexokinase/Glucokinase
reduced to lactate (i.e. anaerobic glycolysis). Accumulation Hexokinase activity is subject to feedback inhibition by
of lactate and consequent fall in pH is potentially haz- glucose 6-phosphate. This prevents intracellular accumu-
ardous and should be prevented. Mechanisms exist in body lation of glucose 6-phosphate.
which prevent such excessive accumulation of lactate. Glucokinase, that specifically effects phosphorylation
The lactate first diffuses out into the blood circulation, of glucose in liver, is an inducible enzyme. Availability of
the plasma membrane of muscle cells being freely per- substrate glucose induces the enzyme synthesis, proba-
meable to lactate. It is carried to liver, and is oxidized to bly through insulin. It is free from feedback inhibition
pyruvate in hepatocytes by the lactate dehydrogenase reac- by glucose 6-phosphate.
tion. Pyruvate, so produced, enters the gluconeogenic
sequence and converted to glucose, which is then trans- Regulation of Phosphofructokinase
ported to skeletal muscles (Fig. 9.7). Most important control point for glycolysis is through regu-
Thus, a cyclic process is set up between liver and mus- lation of phosphofructokinase (PFK1), a complex allosteric
cle, which ensures efficient reutilization of lactate by the enzyme. Its activity is influenced by cellular energy
body. It is referred to as the Cori cycle, named after Carl charge (high energy charge inhibits PFK1 and low energy
BOX 9.3
Rate of Glycolysis is Regulated by Cellular Energy Charge
An overview of regulation of glycolysis shows that its rate is adjusted according to the energy content of the cell. In gen-
eral, when the cellular energy content is low, the catabolic pathways are accelerated so as to provide the required energy;
and conversely, when the cell has sufficient energy, inhibition of these pathways occurs. The anabolic pathways are regu-
lated in a reciprocal manner, i.e. stimulated by high energy content and inhibited by low energy content of the cell.
Evidently, cellular energy content is an important determinant of the rate of various metabolic pathways, including gly-
colysis. It is reflected by the following equation:
ATP
ATP AMP
It is commonly referred to as the energy charge of the cell. Low energy charge signals that the cell needs energy, and
accordingly the catabolic pathways, including glycolysis, are stimulated. When the energy charge is high, inhibition of
glycolysis occurs.
charge stimulates it; Box 9.3), and by a number of posi-

Table 9.3. Allosteric modulators of phosphofructokinase (PFK1)
tive and negative allosteric modulators (Table 9.3):
Activators Inhibitors
1. ATP: ATP is an important inhibitory modulator of the AMP ATP
enzyme activity.
Fructose 2,6-bisphosphate Citrate
When the intracellular ATP level rises (indicating high
ADP Ca2
cellular energy charge), the enzyme activity is inhib-
ited. This is because of binding of ATP to the allo- K Mg2
steric site of the PFK1, which results in diminished Phosphate Low pH
affinity of the enzyme for fructose 6-phosphate.
Conversely, low intracellular ATP level results in
removal of ATP from the allosteric site, which Covalent Modulation of PFK2
results in enhanced activity of the enzyme. Activity of PFK2 is regulated by phosphorylation-dephos-
2. Citrate: The inhibitory effect of ATP on PFK1 is enhanced phorylation mechanism.
by citrate, an intermediate of TCA cycle. Regulation of The dephosphorylated enzyme acts as kinase
glycolysis by an intermediate of citric acid cycle ensures The phosphorylated enzyme acts as phosphatase.
that rates of these two pathways, keep pace with each
other. Whenever TCA cycle is impeded, accumulation The following hdormones regulate interconversion of
of citrate results, which in turn slows down glycolysis the phosphorylate/dephosphorylated forms:
as well. Glucagon: It acts through the second messenger cAMP
3. AMP: Effect of AMP on PFK1 is stimulatory since it (and cAMP activated protein kinase A) and phosphory-
opposes the inhibitory effect of ATP on this enzyme. lates the enzyme. This results in activation of the phospha-
Reciprocal influences of ATP and AMP have additive tase activity of PFK2, causing hydrolysis of fructose
effect on PFK1. This is because when intracellular AMP 2,6-bisphosphate. This decreases activity of PFK1 and,
level is high, ATP level is correspondingly low and therefore, the rate of glycolysis falls. Thus, the overall
both (low ATP and high AMP) stimulate PFK1, thereby effect is:
speeding up the rate of glycolysis.
Glucagon cAMP induced phosphorylation stimula-
4. Fructose 2,6-bisphosphate (F-2,6-BP): This compound is
tion of phosphatase activity of PFK2 concentration of
the most important allosteric modulator (activator) of
fructose 2,6-bisphosphate activity of PFK1, and
PFK1 and, therefore, of glycolysis in liver. It is synthesized
decreased rate of glycolysis.
from fructose 6-phosphate by phosphofructokinase 2
(PFK2), a different enzyme from PFK1, and is hydro- Insulin: opposes glucagon by lowering the cellular cAMP
lyzed back to fructose 6-phosphate by fructose concentration that promotes dephosphorylation of the
2,6-bisphosphatase 2 (FBPase2). Interestingly, both PFK2 enzyme and hence stimulates its kinase activity.
and FBPase2 are activities catalyzed by the same poly- Thus, by adjusting the cellular concentration of fructose
peptide; hence this is a bifunctional enzyme (Fig. 9.8). 2,6-bisphosphate, insulin and glucagon can influence the
cAMP Insulin Feedback

inhibition
+
−
ATP Hexokinase
Phosphofructokinase-2 Glucose Glucose 6-P Fructose 6-P
Glucokinase
ADP ATP
(liver)
Phosphofructokinase
+ AMP, fructose 2,6-P
+
Fructose − ATP, citrate
Fructose 2,6- +
6-phosphate bisphosphate PFK1 Insulin
ADP
Pi
Fructose 1,6-BP
ATP ADP
Fructose 2,6-
Bisphosphatase Phosphoenol-
Pyruvate
pyruvate
Pyruvate kinase
+ ATP, Alanine
(active) −
cAMP Glucagon
Pi Protein
Fig. 9.8. Regulation of phosphofructokinase-2 (PFK2), an kinase
Phosphatase
enzyme having kinase activity and phosphatase activity. The Pyruvate +
+ cAMP Glucagon
kinase-P
phosphatase activity is stimulated by glucagon (via cAMP (inactive)
induced phosphorylation), and the kinase activity by insulin.
Fig. 9.9. Regulation of glycolysis.
glycolytic activity (and gluconeogenic activity, discussed later)
within minutes.
Fructose 6-phosphate is a potent modulator of PFK2- Covalent modulation: Pyruvate kinase is inhibited by the
activity. It not only stimulates the synthesis of F-2,6-BP, cAMP-induced phosphorylation of a serine side chain in
but also inhibits its hydrolysis. F-2,6-BP in turn strongly the enzyme protein (Fig. 9.9). Thus glucagon, which acts
activates PFK, and hence stimulates glycolysis. The over- via cAMP-dependent protein kinase, inhibits the enzyme
all effect is that when fructose 6-phosphate levels are activity (thereby inhibiting hepatic glycolysis); and con-
high, such as after meals, the PFK, (and hence) gly- versely insulin stimulates it.
colysis is stimulated.

Because of various aforementioned regulatory influ- Pyruvate kinase is activated by fructose 1,6-bisphosphate
ences the activity of PFK1 can increase up to 700-fold but allosterically inhibited by ATP and alanine. Like PFK,
from the basal resting level. it is also regulated hormonally by glucagon.

The glycolytic reactions catalyzed by hexokinase, phos-
phofructokinase and pyruvate kinase are metabolically E. Diseases Associated with Glycolysis
irreversible and subject to regulation. Phosphofructokinase
is most important, of which fructose 2,6-bisphosphate is Excessive accumulation of lactic acid occurs in a number of
most important regulator. conditions, described in Chapter 1, to cause lactic acidosis.
It causes severe metabolic acidosis, and is a potentially
lethal condition (Case 9.1).
Regulation of Pyruvate Kinase
Deficiencies of glycolytic enzymes, most commonly
Pyruvate kinase is next in importance to PFK1 in the regu-
of pyruvate kinase and hexokinase, have been reported.
lation of glycolysis. It has three isozyme forms, all cata-
In these rare conditions, the predominant clinical mani-
lyzing the final reaction (Reaction 10) of the glycolytic
festation is haemolytic anaemia because energy depleted
sequence. Its activity is regulated allosterically and by
erythrocytes are easily destroyed. A rare inherited disease
covalent modulation.
causing pyruvate dehydrogenase (PDH) depletion has been
Allosteric regulation: Pyruvate kinase is inhibited allosterically reported (incidence 1 in 250,000 births). Metabolism of
by ATP and alanine and activated by fructose 1,6-bisphos- pyruvate being blocked, lactic acidosis frequently devel-
phate. The concentration of fructose 1,6-bisphosphate is ops. Blockage of the PDH reaction can also occur in beri-
increased whenever PFK1 activity increases, and so its effect on beri (thiamine pyrophosphate being a cofactor in PDH)
pyruvate kinase is an example of feed-forward stimulation. and among alcoholics (Case 18.1).
Aldolase B is an isoenzyme of the aldolase (see Reaction

III. Feeder Pathways 4-glycolysis), and it can cleave both fructose 1,6-
bisphosphate and fructose 1-phosphate. Glyceraldehyde
Glycolysis is not exclusively for the catabolism of glucose; is phosphorylated by the enzyme triokinase to glyceral-
many other carbohydrates enter the glycolytic sequence dehyde 3-phosphate, which along with DHAP, is metab-
in course of their metabolism. The metabolic pathways by olized further by glycolysis or gluconeogenesis.
which various monosaccharides, disaccharides and polysac- Fructose is absorbed from the intestine less rapidly
charides enter the glycolytic sequence are called feeder pathways. than glucose, but once in the blood it is metabolized
Separate feeder pathways exist for various monosaccha- nearly twice as rapidly as glucose. It bypasses the phospho-
rides, disaccharides and polysaccharides. fructokinase step in liver, and therefore, its metabolism is
simpler than that of glucose (Box 9.4).
A. Feeder Pathways for Disorders of Fructose Metabolism
Monosaccharides
Essential fructosuria: It is a rare medical problem
associated with fructose metabolism. The enzyme
Metabolism of Fructose fructokinase is deficient in this disorder and therefore,
D-Fructose is present in free form in many fruits, and
fructose cannot be metabolized as rapidly as in normal
honey. The major dietary source is sucrose (cane sugar),
subjects. Plasma fructose levels rise, and fructose may
a disaccharide consisting of glucose and fructose. In the
appear in urine. The condition is asymptomatic,
body, entry of fructose into the cells is not dependent
sometimes detected incidentally during urinalysis when
on insulin. This is in contrast to glucose, which requires
insulin for this purpose.
ATP ADP
1
The Pathway Fructose Fructose 1-phosphate
Some of the fructose in the cells is phosphorylated to the Fructokinase
glycolytic intermediate fructose 6-phosphate by hexoki- Aldolase-B 2
nase in muscle and adipose tissue. Affinity of hexokinase
for fructose, however, is extremely low (Km approxi-
mately 20 times higher than that for glucose), so this
pathway is normally insignificant, and is important only
when the fructose concentration is very high. Most of the Glyceraldehyde + Dihydroxyacetone
phosphate
fructose is phosphorylated by the enzyme fructokinase in ATP
the liver, kidney, and intestine. The liver accounts for
nearly half of the total fructose metabolism. Fructokinase Triokinase
catalyzes phosphorylation of fructose at C-1 to form ADP

fructose 1-phosphate (Fig. 9.10). The latter is cleaved
Glyceraldehyde 3-phosphate
into dihydroxyacetone phosphate (DHAP) and glyceral-
dehyde by the enzyme aldolase B. Defective action of this Fig. 9.10. Conversion of fructose to intermediates of glycolysis
enzyme leads to a disorder called hereditary fructose in liver. ( = metabolic blocks: 1. in essential fructokinase, 2. in
intolerance (Case 9.2). hereditary fructose intolerance).
BOX 9.4
Is Fructose Useful in Parenteral Nutrition
Metabolism of fructose bypasses the phosphofructokinase step in liver, and therefore simpler than metabolism of glucose.
Because of this, fructose was considered to be more useful than glucose in patients requiring parenteral nutrition. However,
use of fructose in such patients has severe limitation. Fructose is rapidly phosphorylated to fructose 1-phosphate in hepa-
tocytes. This reaction is so fast that a much larger amount of fructose 1-phosphate is generated than can be metabolized.
This ties up the available inorganic phosphates, thereby depleting the liver cell of P1 and ATP. Consequences of such deple-
tion are highly hazardous. Therefore, use of fructose is not recommended in parenteral nutrition.
“reducing sugar” (fructose) is present. Restriction of Hypertriglyceridaemia: Fructose is rapidly metabolized to

dietary fructose is effective to treat this. yield acetyl CoA, which is channeled into lipogenesis
(fatty acids and triglycerides synthesis) in the liver.
Hereditary fructose intolerance: These are disorders caused
by the deficiency of one of the fructose metabolizing Hyperuricaemia: This is due to excessive accumulation of
enzymes. The enzymes involved are: ADP and AMP (due to lack of Pi) followed by their deg-
radation to uric acid.
1. Aldolase B: Deficiency of aldolase B results in intracel-
lular accumulation of fructose 1-phosphate. There is
Metabolism of Galactose
vomiting, jaundice, and hypoglycaemia caused by intra-
D-Galactose is present in milk as one of the monomeric
cellular accumulation of fructose 1-phosphate. This
constituents of lactose; the other one is glucose. Galactose
compound allosterically inhibits liver phosphorylase to
is liberated in the intestine by action of the brush border
block glycogenolysis, thus leading to hypoglycaemia.
enzyme, lactase.
Besides tying up phosphate, and thereby impairing ATP
synthesis, fructose 1-phosphate inhibits aldolase and
The Pathway
phosphohexose isomerase; these changes result in liver
Entry of galactose into peripheral cells is independent of
damage, and jaundice is commonly seen. More about
insulin. Galactose is phosphorylated at C-1 by the enzyme
hereditary fructose intolerance is given in Case 9.2.
galactokinase to form galactose 1-phosphate. Galactose
2. Fructose 1,6-bisphosphatase: Deficiency of this gluco-
1-phosphate is converted to glucose 1-phosphate by the
neogenic enzyme causes fructose intolerance similar
action of galactose 1-phosphate uridyl transferase. In this
to aldolase-B deficiency, but these patients also have
reaction, uridine diphosphate (UDP) acts as a carrier of
fasting hypoglycaemia. They can form glucose from
hexose molecule (Fig. 9.11). The net reaction of the path-
stored glycogen, but gluconeogenesis is blocked.
way is the ATP-dependent conversion of galactose to glu-
Glycogen degradation can maintain a reasonably
cose 1-phosphate. The latter enters glycolytic sequence
normal blood glucose level for many hours, but
via glucose 6-phosphate.
the defect in gluconeogenesis results in dangerous
UDP-galactose is an important intermediate formed in
hypoglycaemia when the period of fasting exceeds
the second reaction (from galactose 1-phosphate and
14–18 hours.
UDP-glucose). It can be epimerized to UDP-glucose, as
shown in the Figure 9.11. The reversibility of epimerization
Inborn errors of fructose metabolism cause hypoglycae- makes it possible to obtain UDP-galactose from UDP-
mia and liver damage. glucose, so that galactose is not an essential nutrient in diet.
UDP-Galactose has synthetic roles, being involved in
the following reactions:
Excess Fructose is Hazardous for Body 1. It is an active donor of galactose residue during syn-
Based on the reasoning that insulin-dependent PFK1
thesis of lactose.
reaction is bypassed by fructose, diabetic diets were for-
mulated in the past with fructose as the predominant car- UDP-Galactose Glucose Lactose UDP
bohydrate. It was soon found, however, that excess 2. Glycolipids, glycosaminoglycans, glycoproteins and cere-
fructose is toxic because it can cause damaged liver and brosides are some other important compounds, which
cause increased lactate formation, hypertriglyceridaemia depend on UDP-galactose as the donor of galactose.
and hyperuricaemia, not only in individuals with diabe-
tes but in normal subjects as well. Galactosaemia
Liver damage: The activity of fructokinase far exceeds that Inborn errors due to deficiency of one of the enzymes of galactose
of aldolase B, so fructose 1-phosphate tends to accumulate metabolism cause galactosaemia. The deficient enzyme may
(concentration in liver can reach up to 10 mol/g). This be galactokinase, galactose 1-phosphate uridyl transferase, or
compound allosterically affects several enzymes of car- rarely epimerase. Deficiency of the uridyl transferase,
bohydrate metabolism and also ties up substantial por- known as classical galactosaemia, is best known.
tion of the phosphate in the cell. This can impair oxidative Classical galactosaemia: This is an autosomal recessive
phosphorylation, and lower the synthesis of ATP from disease with an estimated population incidence of 1 in
ADP, with consequent damage to liver cells. 40,000. Deficiency of the uridyltransferase leads to accumu-
Hyperlactataemia: This can be traced to a rapid metabo- lation of galactose and galactose 1-phosphate after inges-
lism of fructose to pyruvate, which is in equilibrium with tion of milk or food containing galactose . There is elevated
lactate. concentration of galactose in blood (galactosaemia) and
Galactose
ATP
Galactokinase
ADP
Galactose 1-phosphate
UDP-glucose
Galactose 1-phosphate
Epimerase uridyl transferase
UDP-galactose
Glucose 1-phosphate
Phosphoglucomutase
Glucose 6-phosphate Glycolysis
Fig. 9.11. Galactose entering glycolysis via the galactose-glucose interconversion pathway, a four-step reaction sequence.
urinary elimination (galactosuria). Other salient features Diagnosis: Presence of reducing material (galactose) in
are as follows: urine with a negative glucose oxidase suggests diagnosis. In
1. Excess galactose is reduced to galactitol (dulcitol) by addition, galactose 1-phosphate uridyl transferase activity can
aldose reductase, the same enzyme that forms sorbitol be measured in RBCs: the patients completely lack the
from glucose in the polyol pathway. Galactitol, like enzyme, and the unaffected heterozygotes have a reduced
sorbitol, accumulates in lens and causes cataract. enzyme activity.
2. Accumulation of galactose 1-phosphate and concom-
Treatment: The patient is placed on a milk-free diet.
itant depletion of inorganic phosphate causes liver
Synthetic diets like soya milk, which are free of galactose,
dysfunction. It is evident within weeks after birth: feed-
are recommended. The galactosaemic child, even when
ing difficulties and vomiting result in poor weight gain,
sustained on galactose-free diet, thrives well because galac-
and jaundice is present that often is misdiagnosed as
tose needed for the synthesis of biomolecules, can be
physiological jaundice of newborn. Excess galactose
obtained from UDP-glucose by epimerization reaction.
1-phosphate inhibits glycogen phosphorylase and phos-
phoglucomutase; both impede glycogenolysis and glyco- Galactokinase deficiency: Generally, galactokinase-deficient
gen accumulates in liver. patients do not suffer from liver or renal complications,
3. The central nervous system is affected as well, as evi- but develop cataract at a very early age (within 10 months
denced by an abnormal lethargy or irritability. Hypo- after birth).
glycaemia is commonly observed because galactosaemia
provides persistent stimulus for insulin secretion from
the pancreas. Galactose in erythrocytes inhibits glucose
Metabolism of Mannose
D-Mannose is a constituent of various polysaccharides
6-phosphate dehydrogenase and hence HMP-shunt,
and glycoproteins. It is phosphorylated at C-6 by hexoki-
which reduces the supply of NADPH. This results in
nase to form mannose 6-phosphate. The latter is isomer-
haemolysis, as discussed in Chapter 10.
ized by the enzyme phosphomannose isomerase to fructose
4. In severe, untreated cases of galactosaemia, mental
6-phosphate, a glycolytic intermediate.
deficiency, liver cirrhosis, cataract, aminoaciduria and
albuminuria develop.
B. Feeder Pathways for Disaccharides

Classical galactosaemia, caused by deficiency of and Polysaccharides
enzyme galactose 1-phosphate uridyl transferase, results
in accumulation of galactose and galactose 1-phosphate.
The disaccharides are cleaved into the constituent mono-
Diversion of galactose in the galactitol is implicated in
saccharides by disaccharidases such as lactase, maltase,
pathogenesis of cataract.
and sucrase. Lactose is cleaved by lactase into glucose and
galactose, sucrose by sucrase into glucose and fructose, acetyl CoA. A mutase enzyme then converts it to N-acetyl
and maltase cleaves maltose into two glucose molecules. glucosamine 1-phosphate, which reacts with UTP to pro-
The disaccharidases are present in outer surface of the duce the nucleotide activated form, UDP-N-acetyl glucos-
epithelial cell lining of the small intestine. The monosaccha- amine. The latter is epimerized to form UDP-N-acetyl
rides, generated by their action are carried to liver by portal galactosamine (Fig. 9.12).
blood. In liver, the monosaccharides are further metabo- Both the nucleotide activated forms (UDP-N-acetyl
lized through glycolytic sequence, as described earlier. glucosamine and UDP-N-acetyl galactosamine) are then
Brush used for synthesis of complex molecules, e.g. glycopro-
Disaccharides border Monosaccharides teins, proteoglycans and glycolipids. UDP-N-acetyl glu-
enzymes
cosamine can enter an alternate pathway to form NANA.
Maltase Glucose + Glucose
Maltose
Lactase
Lactose Glucose + Galactose Glycolysis
Sucrase
Sucrose Glucose + Fructose IV. Tricarboxylic Acid Cycle
Similarly, the polysaccharides are first hydrolyzed into Tricarboxylic acid (TCA) cycle is also called Krebs cycle
the constituent monosaccharides, which are then funneled or the citric acid cycle. It is a cyclic pathway occurring in
into the central glycolytic sequence. mitochondria, also referred to as the “central catabolic
pathway”. It occupies a central place in catabolism because
Monosaccharides, e.g. fructose, galactose, and man- catabolic pathways involving various biomolecules of
nose, as also the common disaccharides and polysac- diverse origin and chemical nature ultimately terminate in
charides, are enzymatically channeled into the pathway TCA cycle. The end product of these pathways is one of the
of glucose metabolism. intermediates of TCA cycle. For example, the glycogenic
amino acids yield pyruvate, oxaloacetate, or -ketogluta-
Lactose intolerance is a commonly encountered clinical rate; the ketogenic amino acids yield acetyl CoA (which
condition, due to deficiency of the intestinal lactase. Lactose is the main precursor of TCA cycle), and the end product
cannot be digested and is oxidized by bacteria in gut, pro- of degradation of various fatty acids and carbohydrates is
ducing gas, bloating and watery diarrhoea. Deficiency dis- also acetyl CoA (Fig. 9.13). Thus, TCA cycle is spoken of
orders of some other disaccharidases are also known to exist. as the meeting point where catabolic pathways converge.
Metabolism of Amino Sugars

TCA cycle is the final common pathway of the oxidation
The amino sugars are required for the synthesis of glyco-
of the acetyl CoA, formed from carbohydrates, fatty
lipids, glycoproteins, and proteoglycans. They are syn-
acids and amino acids.
thesized from fructose 6-phosphate. The amino group is
derived from amide group of glutamine, the reaction
TCA cycle serves two main functions in metabolism:
being catalyzed by amido transferase.
Glucosamine 6-phosphate forms its acetylated derivative (a) to provide an important pool of metabolic interme-
(N-acetyl glucosamine 6-phosphate) by condensing with diates for synthesis of many useful compounds, and
Acetyl
CoA CoA N-Acetyl N-Acetyl
Fructose Glucosamine glucosamine glucosamine
6-phosphate transferas 6-phosphate 6-phosphate 1-phosphate
ido e
Am
Glutamine Glutamic acid UPT
PPi
UDP-N-Acetyl UDP-N-Acetyl
galactosamine glucosamine
Glycoproteins, proteoglycans
glycolipids, NANA
Fig. 9.12. Metabolism of amino sugars (UDP-N-Acetyl glucosamine and UDP-N-Acetyl galactosamine).
Lipids Carbohydrates Proteins
Fatty acids Glycerol Amino acids

Ketogenic Glucogenic
Pyruvate
Acetyl CoA
Oxaloacetate Citrate
Isocitrate
TCA
α-Ketoglutarate
Fig. 9.13. TCA cycle as the meeting point of various catabolic pathways.
(b) to break down some compounds for generation of Coenzymes: Lipoic acid, FAD, NAD, coenzyme A and
energy. The energy obtained is captured by reducing thiamine pyrophosphate (TPP) are the coenzymes in this
NAD and FAD to NADH and FADH2, respectively. complex. They serve as transient carriers of functional groups
To obtain full usage of the energy generated in this or participate in the oxidation–reduction reactions.
cycle, NADH and FADH2, are processed by another Aggregation of various enzymes in a complex greatly
pathway (oxidative phosphorylation) where their increase the efficiency of the transformation. This is because
energy is converted to ATP. the intermediate products are not released to the medium
but are channeled to the succeeding enzymes of the path-
way. Thus a metabolic intermediate is available for the next
A. Pyruvate Dehydrogenase Complex: reaction immediately after its production. Outline of the
A Bridge Between Glycolysis and reaction sequence and role of various enzymes and coen-
TCA Cycle zymes of this complex are summarized in Figure 9.14.

Acetyl CoA, one of the two major reactants in the first reac- The pyruvate dehydrogenase multienzyme complex, which
tion of TCA cycle (the other being oxaloacetate), is mainly contains three enzymes and five coenzymes, generates
generated from pyruvate. This conversion involves a acetyl CoA from the glycolytic product, pyruvate.
series of complex reactions, catalyzed by a multienzyme
complex, called pyruvate dehydrogenase complex (PDH).
Strictly speaking, this reaction sequence is not a part of
the TCA cycle. It is, however, discussed here along with
B. TCA Cycle as a Cyclic Pathway
TCA cycle, because it serves as a bridge between glycolysis
TCA cycle consists of eight sequential reactions. It is
and TCA cycle.
begins with condensation of a four carbon oxaloacetate
The pyruvate dehydrogenase complex is located in the
(OAA) molecule with an acetyl CoA molecule (2-carbon)
mitochondrial matrix. It is a multimolecular aggregate (MW
to form a six carbon citrate molecule (Reaction 1; Fig.
9 106) consisting of three enzymes and five coenzymes.
9.15). In subsequent reactions of TCA cycle, two carbon
Enzymes: Pyruvate decarboxylase, dihydrolipoyl transacety- atoms are lost from the citrate in the form of CO2
lase and dihydrolipoyl dehydrogenase are the three enzymes (Reactions 3 and 4, Fig. 9.14). A series of modifications
present in the complex. There are about 60 molecules of occur, in the remaining four carbon atoms, in successive
dihydrolipoyltransacetylase and about 20–30 molecules steps, to ultimately form oxaloacetate (OAA). Thus, the
each of the other two enzymes in each complex. last intermediate of one cycle (i.e. OAA) is ready for use
Pyruvate
CH3 1 OH decarboxylase
C O + TPP CH3 C COO–
Thiamine 2
COO–
pyrophosphate TPP
Pyruvate
Condensation
product
TPP CO2
Dihydrolipoyl
O transacetylase
S C CH3
Acetyl lipoic L CH3 C− OH
acid 3
SH
TPP
Hydroxyethyl
CoASH
intermediate
4
O
CH3 C SCoA
Dihydrolipoyl
Acetyl CoA dehydrogenase
SH S
L Lipoic acid
L
SH 5 (oxidized)
S
Lipoic acid FAD FADH2
TCA cycle (reduced)
NADH + H+ NAD+
Fig. 9.14. Mechanism of conversion of pyruvate to acetyl CoA by pyruvate dehydrogenase complex. Step 1: Pyruvate and thia-
mine pyrophosphate (TPP) combine to form a condensation product. Step 2: Pyruvate decarboxylase catalyzes release of carbon
dioxide from the condensation product to form a hydroxyethyl intermediate. The latter is attached to the reactive carbon of the
TPP. Step 3: Transfer of acetyl group from the hydroxyethyl intermediate to the lipoic acid (oxidized) occurs to form acetyl lipoic
acid. The reaction is catalyzed by dihydrolipoyl transacetylase. Step 4: Transfer of an acetyl group from the acetyl lipoic acid to
coenzyme A (CoASH) occurs next, converting the latter to acetyl CoA. The acetyl CoA then enters the TCA cycle. The other prod-
uct of this reaction is reduced lipoic acid. Step 5: This step regenerates the oxidized lipoic acid (from the reduced lipoic acid),
which then participates in the next cycle of reactions. This conversion is catalyzed by the dihydrolipoyl dehydrogenase component
of the enzyme complex, which catalyzes transfer of the reducing equivalents first to FAD and then to NAD.
Overall: Pyruvate NAD CoASH Acetyl CoA CO2 NADH H.
Acetyl CoA mitochondrial membrane (Chapter 14). This arrange-

(2C)
ment ensures that the free energy liberated during reactions
Oxaloacetate Citrate (6C) of the TCA cycle is promptly trapped in the form of ATP.
1
(4C) 2
8
Malate Isocitrate (6C)
(4C) 3
CO2
C. Reactions of TCA Cycle
7 α-Ketoglutarate (5C)
4
Fumarate
6 CO2 The reactions brought about by different enzymes of the
(4C) 5 Succinyl CoA
Succinate TCA cycle are depicted in Figure 9.16. All the enzymes
(4C)
(4C) are present in mitochondrial matrix except succinate
dehydrogenase which is located in the inner mitochon-
Fig. 9.15. TCA cycle: Oxaloacetate (OAA), the last interme-
drial membrane.
diate is also a reactant in the first step. The eight reactions of
the pathway are numbered 1 to 8.
Reaction 1: Synthesis of Citrate from Acetyl CoA and
as a substrate in the next cycle. In this way, there is no net Oxaloacetate
generation of OAA, or of any of the cycle intermediates. Citrate is produced by condensation of acetyl CoA with
The reactions of TCA cycle take place in the mitochon- oxaloacetate in an irreverssible reaction catalyzed by the
drial matrix. This is in close proximity to the reactions of enzyme, citrate synthase. The reaction equilibrium lies far
oxidative phosphorylation, which occur in the inner towards the right (i.e. towards the formation of citrate).
CoASH
O a-Ketoglutarate
C S CoA
dehydrogenase NAD+ 4
complex Mg2+
CH3 Acetyl-CoA NADH + H+
+ CO2
O
C COO− COO−
CH2 CH2
H2O COO− Oxaloacetate CH2

Succinyl-CoA
C S CoA
Citrate
synthase CoASH + H+ O
COO−
GDP + Pi
1 CH2 Succinyl-CoA 5
HO C COO− synthetase Mg2+
GTP
CH2
Citrate
COO− CoASH
COO−
+H2O −H2O
CH2
COO−
CH2
CH2
cis-Aconitate COO− Succinate
Aconitase
C COO− (enzyme-bound)
Fe2+ Succinate
E-FAD
CH 6
dehydrogenase
2 COO− E-FADH2
COO−
−H2O +H2O CH
COO− HC
CH2 COO− Fumarate
H C COO− Fumarase −H2O +H2O
HO C H
COO− 7
Isocitrate
COO
HO C H
NAD+ (NADP+)
CH2
Isocitrate Malate
dehydrogenase COO−
NADH (NADPH) + H+
Mg2+ NAD+
Malate
8
dehydrogenase
3 CO2 NADH + H+
COO−
COO−
CH2
C O
CH2
CH2
C O
α-Ketoglutarate COO− Oxaloacetate
COO−
Fig. 9.16. Reactions of tricarboxylic acid (Krebs) cycle.

TCA cycle derives its other name (citric acid cycle) from acetyl CoA carboxylase, the rate-limiting enzyme of fatty acid
this first intermediate, citrate. However, citrate may leave synthesis. Thus citrate enhances fatty acid synthesis by:
the citric acid cycle to participate in other metabolic Providing the substrate (i.e. acetyl CoA).
pathways as well (Fig. 9.17). Excessive citrate crosses the Stimulating the key lipogenic enzyme (i.e. acetyl CoA
inner mitochondrial membrane through specific tricar- carboxylase).
boxylate carriers and reaches the cytosol, where it provides
acetyl CoA: In the cytosol, citrate has another important regula-
Citrate lyase tory role to play. It adjusts the rate of glycolysis to that of
Citrate Acetyl CoA + OAA
TCA cycle. It does so by inhibiting phosphofructokinase
In cytosol, acetyl CoA serves as a precursor for fatty (PFK1) the rate-limiting enzyme of glycolysis (Table 9.3).
acid synthesis (lipogenesis). Moreover, citrate stimulates Inhibition of this enzyme decreases the rate of glycolysis,
PFK1 Fatty acid

The mechanism of this conversion is similar to that of
the conversion of pyruvate to acetyl CoA (Fig. 9.14). The
same set of coenzymes, i.e. thiamine pyrophosphate,
−
Malonyl CoA lipoic acid, FAD, NAD, and CoASH, is used. Each of
these performs a function analogous to that performed
+ E
in the pyruvate dehydrogenase complex.
Citrate Citrate Acetyl CoA The reaction releases the second CO2 molecule of the
TCA
Cycle cycle and produces the second NADH. The equilibrium
OAA
of the reaction lies towards the right, i.e. towards succinyl
Mitochondrial CoA formation.
matrix
IMM Cytosol
Reaction 5: Cleavage of Succinyl CoA
The enzyme succinyl CoA synthetase (also called succinate
thiokinase) cleaves the high-energy thioester bond in suc-
Fig. 9.17. Citrate, a TCA intermediate, plays an important cinyl CoA (G0‘ is –8.0 kcal/mole) to release large
regulatory role in lipogenesis and glycolysis by modulation of amount of free energy, which is used to produce a GTP
activities of the enzymes, phosphofructokinase (PFK1) and acetyl molecule.
CoA carboxylase (E) (IMM inner mitochondrial membrane). This reaction provides an example of substrate level
phosphorylation, since the production of a high-energy
and therefore, the production of acetyl CoA falls. This phosphate (e.g. GTP) is coupled with enzymatic transfor-
results in decreased rate of TCA cycle since acetyl CoA is mation of a substrate molecule. GTP can produce ATP by
needed in the first reaction of TCA cycle. In this way, rates action of the enzyme nucleoside diphosphokinase.
of glycolysis and TCA keep pace with each other. GTP ADP GDP ATP
Thus, when citrate concentration is high, implying that
the cell is adequately supplied with fuel molecules and, Note: Succinyl CoA may also serve as a substrate for
therefore, further production of energy is not required, the haem synthesis.
energy-yielding catabolic pathways (e.g. glycolysis and
Reaction 6: Oxidation of Succinate
TCA cycle) are inhibited. The biosynthetic pathway (i.e.
A pair of reducing equivalents is removed from succinate
fatty acid synthesis) is favoured at the same time. This
by the enzyme succinate dehydrogenase to form fumarate.
illustrates a fundamental principle of biochemistry, i.e. the
FAD serves as a coenzyme in this reaction. Unlike the
catabolic and the anabolic pathways are regulated recipro-
other enzymes of the TCA cycle, which are located in the
cally. If one pathway is favoured, the other is inhibited.
mitochondrial matrix, succinate dehydrogenase is anchored
Reaction 2: Isomerization of Citrate in the inner mitochondrial membrane. It catalyzes removal of
Isomerization of citrate by the enzyme aconitase yields a hydrogen pair from succinate and transfers it to FAD
isocitrate. During this reaction, a transient enzyme- which becomes FADH2. This in turn transfers its electrons
bound intermediate, cis-aconitate is formed. to ubiquinone, which becomes ubiquinol, and is then
transferred to complex III for oxidation (Chapter 14).
Reaction 3: Oxidative Decarboxylation of Isocitrate
The enzyme isocitrate dehydrogenase (IDH) catalyzes Reaction 7: Hydration of Fumarate
removal of two hydrogen atoms from isocitrate (i.e. oxi- Addition of a water molecule to fumarate forms malate.
dation) with a concomitant release of a CO2 molecule It is a reversible reaction, catalyzed by the enzyme
(i.e. decarboxylation). -Ketoglutarate is the reaction fumarase.
product.
NAD serves as a coenzyme in this step, and is con- Note: In spite of being reversible, this reaction always
verted to NADH by accepting a pair of hydrogen atoms. proceeds unidirectionally, i.e. towards formation of malate.
NADP can also serve as a coenzyme in this reaction. This is because of the thermodynamic pull in TCA cycle
Isocitrate dehydrogenase is one of the few enzymes that are (discussed later).
capable of using both NAD and NADP as coenzymes.
Reaction 8: Oxidation of Malate
Reaction 4: Oxidative Decarboxylation of -Ketoglutarate Removal of a pair of reducing equivalents from malate
Like the previous step, this one also involves oxidation by the enzyme malate dehydrogenase produces oxaloace-
and decarboxylation. The reaction is catalyzed by the tate. The reaction generates the third NADH molecule of
enzyme -ketoglutarate dehydrogenase, to produce succi- the cycle. Malate to oxaloacetate conversion is the last
nyl CoA. reaction of the cycle. Oxaloacetate, generated in this step,
BOX 9.5
Inhibitors of TCA Cycle
TCA cycle is inhibited by a number of chemicals, which include the following:
Fluoroacetate: It inhibits the enzyme aconitase (non-competitively).
Arsenite: It causes non-competitive inhibition of -ketoglutarate dehydrogenase.
Malonate: It causes competitive inhibition of succinate dehydrogenase.
condenses with another molecule of acetyl CoA to initi- Glucose

ate another cycle.
Glycolysis
The following equation summarizes all the reactions 2 NADH 6 ATP
of the TCA cycle.
2 ATP
Acetyl CoA 3NAD FAD GDP Pi 2H2O
2 Pyruvate
2CO2 3NADH 2H FADH2 GTP CoA-SH
Pyruvate 2 NADH 6 ATP
Several chemicals can inhibit reactions of the TCA dehydrogenase
cycle, as outlined in Box 9.5.
2 Acetyl CoA

A four-carbon compound oxaloacetate reacts with acetyl 6 NADH 18 ATP
TCA cycle
2 FADH2 4 ATP
CoA to form six-carbon citrate, which is then converted back 2 GTP 2 ATP
to oxaloacetate in the remaining reactions of the TCA cycle.
38 ATP
The following points about TCA cycle are noteworthy: 4 CO2
1. Two carbon atoms enter the cycle as acetyl CoA (and Fig. 9.18. Energetics of glucose oxidation. 38 ATPs are gen-
condense with oxaloacetate) and two carbons leave in erated by complete oxidation of glucose via glycolysis, PDH
the form of two molecules of CO2. Thus, TCA basically reaction, and TCA cycle.
involves oxidation of acetyl CoA to carbon dioxide
NADH in this manner generates three ATPs, whereas two
and, as such there is no net consumption or regeneration
ATPs are produced from one FADH2 molecule. In addition,
of oxaloacetate or any of the other cycle intermediates.
one GTP is produced during enzymatic transformation
2. Some of the reactions of TCA cycle are reversible and yet
of succinyl CoA (Reaction 5), which generates an ATP
they always proceed unidirectionally. This is because
through action of nucleoside diphosphokinase. Hence, each
equilibrium of some of the reactions of TCA cycle
cycle of TCA cycle, produces 12 ATP molecules.
(e.g. Reactions 1, 3 and 4, Fig. 9.12) lies far towards
the right. These reactions are irreversible and have 3 NADH (Reaction 3,4 and 8) 9 ATP
strong tendency to proceed unidirectionally, towards 1 FADH2 (Reaction 6) 2 ATP
the right. This generates a strong “thermodynamic pull“ so 1 GTP (Reaction 5) 1 ATP
that rest of the (reversible) reactions are also pulled TOTAL (Each cycle) 12 ATP
in that direction. As mentioned earlier, fumarate to
malate conversion which is freely reversible other- However, two acetyl CoA molecules are generated from
wise, always proceeds in direction of malate forma- each glucose, therefore, this cycle occurs twice, generating
tion because of the thermodynamic pull. 6NADH, 2FADH2 and 2GTPs (Fig. 9.18) Hence , total 24
ATPs are generated. Thus, complete oxidation of glucose
via glycolysis, pyruvate dehydrogenase, the Krebs cycle and the
D. Energy Yield from TCA Cycle oxidative phosphorylation pathway yields 38 ATPs. Since
two ATPs were initially used during stage I reactions of gly-
There are four dehydrogenation reactions in TCA cycle colysis, the net yield is 36 ATPs. Compared with anaerobic
(Reactions 3, 4, 6 and 8), which generate three NADH and pathway, the oxidative pathway thus yields 18 times more
one FADH2 molecules. These reduced coenzymes donate energy in the form of ATP. However, this is still far less than
electrons to the electron transport chain and generate the energy obtained by burning of a molecule of glucose in
ATPs by oxidative phosphorylation. Oxidation of each calorimeter (2780 kJ), which is sufficient to generate about
91 ATPs (one ATP has energy bond equivalent to 30.5 kJ). Aspartate OAA
Evidently, only about 40% of the energy locked in chemical bonds Citrate Fatty acids
of glucose molecule is captured for ATP synthesis in our body. Asparagine
a-Ketoglutarate Glutamate
Gluconeogenesis
Pasteur Effect
It has been observed that under anaerobic condition a Succinyl
CoA Glutamine
tissue or microorganism utilizes much more glucose than
it does under aerobic conditions. This is the Pasteur effect, Haem
GABA
and it reflects inhibition of glycolysis by oxygen. It is also porphyrins
observed that the levels of glycolytic intermediates from
Fig. 9.19. Synthetic functions of TCA cycle.
fructose 1,6-bisphosphate onwards decrease while the ear-
lier intermediates increase. Evidently, the Pasteur effect is
because of inhibition of the enzyme phosphofructokinase. which in turn are required for the synthesis of other non-
The inhibitory effect is caused by citrate and ATP, the essential amino acids (Chapter 13), as also of purines
compounds produced in presence of oxygen. and pyrimidines.
Others
Crabtree Effect Succinyl CoA can be diverted from TCA cycle for biosyn-
When oxygen supply is kept constant and glucose con- thesis of porphyrins and haem (for further information
centration is increased, the oxygen consumption by cells refer to Box 9.6). The -ketoglutarate can form gluta-
falls. This is called Crabtree effect, which is basically opposite mate, from which -aminobutyric acid (GABA), an
of Pasteur effect. It is seen in the cells that have a high rate inhibitory neuro-transmitter is produced.
of aerobic glycolysis. In such cells the glycolytic sequence
consumes much of the available Pi and NAD, which
limits their availability for oxidative phosphorylation. As Some TCA intermediates are substrates for gluconeo-
a result, rate of oxidative phosphorylation decreases, and genesis, amino acid synthesis, and fatty acid synthesis.
oxygen consumption also shows a corresponding fall. Anaplerotic reactions such as the pyruvate carboxylase
reactions replenish TCA cycle intermediates.
E. Synthetic Functions of the TCA Cycle

F. Regulation of TCA Cycle
TCA is not only the final oxidative pathway for various
biomolecules, its intermediates are also substrates for vari- A number of modulators regulate activities of various
ous biosynthetic pathways (Fig. 9.19). Thus, TCA is both enzymes of TCA cycle. Two types of regulatory mecha-
catabolic and anabolic in nature, and hence regarded as nisms—allosteric modulation and covalent modulation
amphibolic. of the enzyme activities operate. Regulation of the pyru-
Some important synthetic pathways connected with vate dehydrogenase complex, the bridge between glycolysis
TCA are as follows: and TCA cycle, and the regulation of the cyclic pathway
are discussed below.
Synthesis of Glucose
Intermediates of TCA cycle can serve as substrates for Regulation of the Pyruvate Dehydrogenase
gluconeogenesis, the pathway that produces glucose Complex
from non-carbohydrate sources. As glucose is synthe- Activity of the pyruvate dehydrogenase complex (PDH) is
sized in this manner some intermediates of TCA cycle are switched-on or switched-off based on cellular energy
removed from the cycle. These intermediates may be sub- needs. When the cellular energy charge is high and sur-
sequently replenished by anaplerotic reactions. plus fuel molecules are present intracellularly, inhibition
of activity of the enzyme complex occurs. Two mecha-
Synthesis of Fatty Acids nisms of regulation have been recognized.
Citrate is used for fatty acid synthesis (Fig. 9.17). Allosteric regulation: The enzyme complex is subject to
allosteric inhibition by the reaction products: acetyl CoA
Synthesis of Amino Acids and NADH. It is stimulated by AMP.
Oxaloacetate and -ketoglutarate can form aspartate and Elevated intracellular concentrations of both acetyl CoA
glutamate, respectively by transamination reactions, and NADH indicate surplus intracellular fuel molecules
BOX 9.6
Anaplerotic (‘Filling-up’) Reactions
The TCA cycle intermediates may be “siphoned off” for various biosynthetic processes, which may deplete the important
pool of these metabolites. For example, removal of succinyl CoA for haem synthesis could gradually deplete intramitochon-
drial concentration of succinyl CoA and hence, that of the other cycle intermediates. If the cycle intermediates were not
replenished, TCA cycle would cease to function. Anaplerotic reactions provide the TCA cycle with metabolic intermediates,
maintaining the activity of the cycle and ensuring that the cycle is never stalled because of lack of these metabolites. Pyruvate
carboxylase (PC) reaction is the classical example of the anaplerotic reaction. PC converts pyruvate to oxaloacetate, which
is required for the initiation of the cycle. Some other important anaplerotic reactions are as below:
Conversion of pyruvate to malate by the cytoplasmic malic-enzyme. Malate can then enter the mitochondrion as a
substrate for the TCA cycle.
Pyruvate may react with aspartate or glutamate in transaminase reactions, producing the TCA cycle intermediates
oxaloacetate and -ketoglutarate, respectively.
Several glycogenic amino acids may serve as source of TCA cycle intermediates (Chapter 13).
NADH : NAD+
Acetyl CoA : CoA
ADP
+ PDH P
Inactive H2O
Pyruvate phosphorylated form
Pyruvate
− dehydrogenase
kinase Protein phosphatase
+ Pi
PDH OH
ATP
Active
Low energy dephosphorylated form
charge-↑ATP : ADP
Fig. 9.20. Regulation of pyruvate dehydrogenase complex (PDH).
and excessive cellular energy charge. Inhibition of the PDH, 1. Pyruvate dehydrogenase kinase: This enzyme phosphory-
under these circumstances, stops further production of lates the enzyme complex, thereby inactivating it.
energy and fuel molecules. Increased activity of the kinase is, therefore, associated
with decreased activity of PDH. Conversely, inhibition
−
NADH + H+
of the kinase activates the enzyme complex (Fig. 9.20).
NAD+
Pyruvate dehydrogenase kinase is allosterically activated
PDH
Pyruvate Acetyl CoA by high energy charge, high [NADH] : [NAD] ratio, and
high [acetyl CoA] : [CoA] ratio and is inhibited by pyru-
vate. Conversely the enzyme activity is inhibited by low
Covalent modulation: The PDH exists in two forms:
energy charge, reflected by elevated ratio of ADP : ATP,
Phosphorylated form, which is inactive.
which in turn results in activation of the PDH. The ele-
Dephosphorylated form, which is active.
vated ADP : ATP ratio signals increased demand for energy
Thus, covalent attachment of a phosphate group ren- production, which is met through activation of the PDH.
ders the PDH complex inactive, whereas removal of this 2. Protein phosphatase: This enzyme removes a phos-
group results in its activation. The following enzymes are phate group from the phosphorylated form of the
involved in interconversion of the phosphorylated- PDH complex to produce the active dephosphory-
dephosphorylated forms. lated form (Fig. 9.20). Thus, it activates the PDH.
Glycolysis modulators of the enzyme. Raised levels of these

metabolites indicate that the cell has adequate supply
Pyruvate of fuel and energy. Under these circumstances, TCA
cycle, which is an energy-yielding pathway, is inhib-
ited so that there is no further production of energy.
Acetyl CoA
− Succinyl CoA
NADH
Regulation of Oxidative Decarboxylation of
1 Citrate Fatty acyl CoA
Isocitrate
Oxaloacetate The enzyme catalyzing this step, isocitrate dehydrogenase,
is activated by ADP and inhibited by high energy charge
Isocitrate NADH : NAD+ and high [NADH] : [NAD] ratio. Elevated levels of
−
3 ADP mitochondrial ADP indicate low energy state of the cell
+
Malate and signals a need for generation of more high-energy
α-Ketoglutarate phosphate molecules (i.e. ATP). Stimulation by ADP
4 accelerates the cycle, generating the required ATP.
Fumarate
−
Succinate
Succinyl CoA NADH Regulation of Oxidative Decarboxylation of
␣-ketoglutarate
Fig. 9.21. Regulation of TCA cycle ( − inhibits reaction, The enzyme catalyzing this step, a-ketoglutarate dehydro-
+ stimulates reaction, numbers 1, 3 and 4 are steps at genase, is allosterically inhibited by its own products
which regulation occurs). (succinyl CoA and NADH) and by high energy charge.
It is evident that low energy charge stimulates TCA so
as to generate more energy; conversely, high energy
Entry of acetyl CoA into the TCA cycle is regulated at the charge inhibits it. The other energy-yielding pathways
pyruvate dehydrogenase step by product inhibition (by
discussed so far, i.e. glycolysis and oxidative decarboxyl-
NADH and acetyl CoA) and by covalent modification.
ation of pyruvate are also regulated similarly.
Regulation of the Cyclic Pathway (Fig. 9.21)

The TCA cycle is regulated at the steps catalyzed by citrate
It occurs at the following steps:
synthase, isocitrate dehydrogenase and a-ketoglutarate
1. Synthesis of citrate (Reaction 1). dehydrogenase. Regulation is accomplished mainly by
2. Oxidative decarboxylation of isocitrate (Reaction 3). substrate availability, allosteric modulation, and feed-
3. Oxidative decarboxylation of -ketoglutarate back inhibition.
(Reaction 4).
Under normal conditions, the rates of glycolysis and
Regulation of Citrate Synthesis TCA cycle are integrated with each other so that only that
This is the most important regulatory step of the path- much glucose is metabolized as required to supply the
way. As in case of most metabolic pathways, where an initial substrate, i.e. the acetyl CoA for the TCA cycle. Rates
initial step is the rate setting step for the pathway as a of these two pathways are matched because of inhibition
whole, the first step of TCA cycle is also most important of phosphofructokinase (PFK1), the key regulatory enzyme of
in regulation. Rate of this reaction is determined by the glycolysis, by a TCA cycle intermediate, citrate. ATP and
following factors: NADH, produced during TCA cycle also inhibit glycoly-
sis by acting as negative modulators of this enzyme.
1. Concentration of the substrate molecules (i.e. oxaloace-
tate and acetyl CoA): Concentration of oxaloacetate
is the most critical factor in determining the reaction G. Glyoxylate Cycle
rate. Concentration of acetyl CoA is also an impor-
tant determinant for pushing the reaction forward. Synthesis of carbohydrate from fat is not possible in the
2. Allosteric modulation of the enzyme: Activity of citrate mammalian cell due to irreversibility of the pyruvate dehy-
synthase, the enzyme catalyzing this step, is mainly drogenase reaction and lack of alternate pathways. How-
regulated by succinyl CoA, which inhibits the enzyme ever, plant cells and some microorganisms can accomplish
activity by decreasing its affinity for acetyl CoA. Fatty this conversion because of a cyclic pathway, termed gly-
acyl CoA and NADH also act as negative allosteric oxylate cycle (Fig. 9.22). It shares several reactions with
gluconeogenesis. The pathway is important because

Fatty acid
some organs, like the brain and the erythrocytes, depend
exclusively on glucose for their energy needs. The brain is
Acetyl CoA said to be a voracious eater of glucose, consuming about
120 grams per day (out of about 160 grams needed by
the entire body) and requires a blood glucose concen-
Oxaloacetate Citrate tration of 70–100 mg/dL. Though degradation of stored
glycogen can also provide glucose, gluconeogenesis is
Acetyl CoA Glyoxylate the only source of glucose during prolonged fasting and
starvation.
Malate Isocitrate
E2 E1 Gluconeogenesis occurs mainly in the liver, and to a
Glucose Succinate lesser extent in renal cortex. On weight basis, the gluco-
α-Ketoglutarate neogenic capacity of the renal cortex equals that of the
Fumarate
liver, but because of the size difference between these tis-
Succinyl sues, the gluconeogenic activity in the kidney amounts
Succinate
CoA to only 10% of that in the liver. However, in certain
abnormal metabolic states, such as prolonged starvation,
Fig. 9.22. Reactions of glyoxylate cycle (shaded area)
kidneys become the major glucose-producing organ.
(E1 isocitrate lyase, E2 malate synthase).
The gluconeogenic pathway is located mainly in
TCA cycle and is considered an anabolic variant of the cytosol, although some precursors are produced in the
latter. It is useful in germinating seeds where the stored mitochondria.
fat is converted to glucose for meeting the energy needs
of the cell. It operates in glyoxysomes, the specialized
cellular organelles where fatty acid oxidation also occurs.
A. Gluconeogenesis: Not a Reversal
of Glycolysis
Reactions
The initial few reactions up to isocitrate formation, are Though most of the reactions of the gluconeogenic
common with TCA cycle, but the isocitrate bypasses the sequence (7 out of 10) are reversal of those of glycolysis
TCA cycle and is cleaved by the enzyme isocitrate lyase to the following three reactions are unique to gluconeogen-
succinate and glyoxylate. The latter compound forms esis, and are catalyzed by a different set of enzymes. They
malate (by condensing with another molecule of acetyl are called the bypass steps:
CoA), which is an intermediate of the TCA cycle. Net result First bypass Conversion of pyruvate to phosphoenolpy-
of these reactions is conversion of two 2-carbon fragments ruvate (PEP).
of acetyl CoA to malate, which is a substrate for gluconeo- Second bypass Conversion of fructose 1, 6-bisphosphate
genesis, and can also enter TCA cycle. to fructose 6-phosphate.
In addition to glyoxylate, the other product of cleavage Third bypass Conversion of glucose 6-phosphate to
of isocitrate is succinate, which is converted to glucose glucose.
via reactions of gluconeogenesis, thereby ensuring net The corresponding steps in glycolysis are formation of
synthesis of carbohydrate from fat. pyruvate from phosphoenol pyruvate (for first bypass);
fructose 1,6-bisphosphate from fructose 6-phosphate
(for second bypass) and glucose 6-phosphate from glu-
The glyoxylate pathway, which operates only in plants,
cose (for third bypass). Recall that only these three reac-
is a variation of TCA cycle. It permits net synthesis of
tions in glycolysis are irreversible. Therefore, an alternate
glucose from acetyl CoA.
set of reactions, unique to gluconeogenesis, is needed to
circumvent these three irreversible glycolytic reactions
(Fig. 9.23). Apart from these three steps, rest of the seven
V. Gluconeogenesis steps of the glycolytic sequence are reversible and are
operative in the gluconeogenesis as well.
Synthesis of glucose from compounds other than carbohydrates
is called gluconeogenesis (i.e. formation of new sugar).

Three reactions are unique to gluconeogenesis, which cor-
A variety of biomolecules such as lactate, pyruvate, gly-
respond to the three irreversible steps of glycolysis. Rest
cerol (derived from triacylglycerols) and -keto acids
of reactions are reversible and common to both pathways.
(derived from amino acid catabolism) are substrates for
Pi Glucose
THIRD BYPASS
Glucose 6-phosphate
Pi Fructose 6-phosphate
SECOND BYPASS
Fructose 1,6-bisphosphate
Glyceraldehyde Dihydroxyacetone
3-phosphate phosphate
G G
L 1,3-Bisphosphoglycerate
L
U Y
C C
O O
N 3-Phosphoglycerate
E L
O Y
G S
E I
2-Phosphoglycerate
N S
E
S
I
S P
GT Phosphoenolpyruvate
Oxaloacetate
FIRST BYPASS
AT
P Pyruvate
Fig. 9.23. The opposing pathways of glycolysis and gluconeogenesis. Steps made in colour are unique to gluconeogenesis.
B. The Bypass Reactions of 1. Transport of pyruvate into mitochondrial matrix: Pyruvate

moves across the inner mitochondrial membrane
Gluconeogenesis (IMM) to enter the mitochondrial matrix through
mediation of specific transport proteins, called the
First Bypass: Conversion of Pyruvate to monocarboxylate carriers monocarboxylate carrier
Phosphoenolpyruvate (PEP) monocarboxylate carriers (MC).
During conversion of phosphoenolpyruvate to pyruvate 2. Pyruvate to oxaloacetate conversion: Pyruvate, a 3-carbon
in glycolytic sequence, release of a large amount of free molecule is converted to 4-carbon oxaloacetate by car-
energy (G0‘–14.8 kcal/mole) occurs under standard con- boxylation reaction catalyzed by pyruvate carboxylase
ditions. For the reverse reaction (i.e. conversion of pyruvate (E1), a biotin-dependent mitochondrial enzyme.
to PEP during gluconeogenesis), input of an equivalent
amount of free energy is required. This is a thermody- Pyruvate CO2 H2O ATP Oxaloacetate
namically unfavourable situation, which is circumvented ADP Pi
by making this conversion occur in two distinct reactions: 3. Transport of oxaloacetate into cytosol: Rest of the enzymes
of gluconeogenesis are located in the cytosol. There-
Conversion of pyruvate to oxaloacetate: 1 ATP
fore, the mitochondrial oxaloacetate, produced in the
hydrolyzed
step 2, must be transported out of the mitochondria
Conversion of oxaloacetate to PEP: 1 GTP hydrolyzed
into cytosol. However, oxaloacetate cannot directly
Sum: Pyruvate to PEP: 1 ATP 1 GTP are hydrolyzed. cross the IMM because the latter is impermeable to
most molecules except for the ones for which specific
The conversion involves the following four steps, carrier proteins exist (such as monocarboxylate carrier
which occurs partly in cytosol and partly in mitochondrial for pyruvate, dicarboxylate carrier for malate, and tricar-
matrix (Fig. 9.24): boxylate carrier for citrate). Therefore, the oxaloacetate
3. Transport of oxaloacetate
into cytosol
DC
Malate Malate NAD+
NAD+
E2
H+ + NADH E2
Oxaloacetate
E3 NADH + H+
GTP
CO2
GDP
Oxaloacetate
4. Conversion of Phosphoenolpyruvate
oxaloacetate to E1 ADP
phosphoenolpyruvate
CO2
ATP
MC 2. Pyruvate to
Pyruvate Pyruvate oxaloacetate
conversion
1. Transport of
pyruvate into
Cytosol mitochondrial
Mitochondrial
matrix
matrix
Inner mitochondrial membrane
Fig. 9.24. Synthesis of phosphoenolpyruvate from pyruvate. The conversion occurs partly in cytosol, partly in mitochondria and
in four steps (MC monocarboxylate carrier, DC dicarboxylate carrier, E1 pyruvate carboxylase, E2 malate dehydrogenase,
E3 phosphoenolpyruvate carboxy kinase).
is converted to malate, a molecule which is capable of A series of reversible reactions follow next, which are
crossing the IMM. The enzyme malate dehydrogenase shared by gluconeogenesis and glycolysis. These reactions
(E2) catalyzes this reaction. result in conversion of PEP to fructose 1,6-bisphosphate
Oxaloacetate NADH H Malate NAD (Fig. 9.23).
Malate is carried out of the mitochondria into cytosol by Second Bypass: Dephosphorylation of
the dicarboxylate carrier (DC). In cytosol, oxaloacetate is
recovered from malate by reversal of the above reaction.
Fructose 1,6-bisphosphate
Hydrolysis of fructose 1,6-bisphosphate by fructose 1,6-
4. Conversion of oxaloacetate to phosphoenolpyruvate: In cyto-
bisphosphatase is the second reaction that is unique to
sol, the oxaloacetate undergoes decarboxylation and
gluconeogenesis. It bypasses the irreversible phosphofructo-
phosphorylation to yield phosphoenolpyruvate (PEP).
kinase reaction. It also provides an energetically favourable
pathway for the formation of fructose 6-phosphate (G0‘
Oxaloacetate GTP PEP CO2 GDP
–3.9 kcal/mole). This reaction is an important control
This reaction is catalyzed by the enzyme phosphoenol- point for gluconeogenesis, discussed later in this chapter.
pyruvate carboxykinase (PEPCK), and is driven forward
by hydrolysis of GTP. Third Bypass: Hydrolysis of Glucose
Thus, a concerted action of several enzymes and car- 6-phosphate
rier proteins permits conversion of PEP to pyruvate; a Glucose 6-phosphatase catalyzes hydrolytic cleavage of the
conversion which is thermodynamically unfavourable phosphate ester to liberate free glucose. Unlike the other
otherwise. Energy of two high-energy phosphate bonds gluconeogenic enzymes, which are cytoplasmic (except
(equivalent to 14.6 kcal/mole), one each from ATP and pyruvate carboxylase), this enzyme resides on the luminal
GTP, are required to drive the conversion across the surface of the endoplasmic reticulum (ER) membrane.
energy “road-block“. The overall equation for this set of Like the fructose 1,6-bisphosphatase reaction, this step
reactions is as below: is also irreversible, and so provides an energetically
Pyruvate ATP GTP PEP ADP GDP Pi favourable pathway for the formation of free glucose
G0‘ 0.2 kcal/mole (G0‘ 2.9 kcal/mole).
yield glucose via the gluconeogenic sequence, as dis-

Gluconeogenesis, the only source of glucose during cussed here.
long-term fasting, is based on a reversal of glycolysis
with specific enzymes bypassing the three irreversible Lactate
reactions of glycolysis. Lactate produced in the skeletal muscles is the major
substrate for gluconeogenesis (Cori’s cycle). It is converted
to pyruvate (in liver) by the lactate dehydrogenase reac-
C. Reversible Steps of Gluconeogenesis tion, which then enters gluconeogenic sequence.
In addition to the three bypass steps, there are seven

reversible reactions common to both these pathways
Amino Acids
Glycogenic amino acids (all except lysine and leucine)
(Fig. 9.23). During gluconeogenesis, their equilibrium is
are catabolized to pyruvate or some intermediate of
pushed in favour of glucose synthesis.
citric acid cycle, which can generate glucose (Chapter
A summary of all reactions of gluconeogenesis, includ-
13). During fasting and starvation, the glycogenic amino
ing the reversible ones is as below:
acids become most important precursors for gluconeo-
2 Pyruvate 4ATP 2GTP 2NADH 2H 4H2O genesis.
Glucose 2NAD 4ADP 2GDP 6Pi 6H
G0‘ 9 kcal/mole
Glycerol
Glycerol is obtained during degradation of adipose triac-
D. Gluconeogenesis is an Expensive ylglycerols. It enters the gluconeogenic sequence in the
Process liver. First, it reacts with ATP to form glycerol 3-phosphate,
which is oxidized to dihydroxyacetone phosphate. The
Several reactions of gluconeogenesis are energetically latter being a glycolytic intermediate can be converted to
unfavourable. It requires input of considerable free energy glucose.
to drive these “uphill” reactions: Thus six phosphoanhy-
dride bonds are required for the synthesis of one glucose Propionate
molecule from two pyruvate molecules (Table 9.4). Catabolism of some amino acids (methionine, isoleu-
Pyruvate carboxylase reactions consume two ATPs, cine) and odd chain fatty acids yields propionyl CoA,
PEP-carboxykinase consume two GTPs, and phosphoglycerate which enters a reaction sequence to ultimately yield suc-
kinase consume two ATPs in the reversal of the substrate cinyl CoA (Chapter 11). Being a TCA cycle intermediate,
level phosphorylation (see reaction 7; Fig. 9.3). succinyl CoA is convertible to glucose.
In addition, two NADH molecules are used up in the gly Fats cannot generate glucose: It is important to note that
ceraldehyde 3-phosphate dehydrogenase reaction, for each mol- fatty acids are NOT substrates for gluconeogenesis, so
ecule of glucose synthesized. Since each NADH can generate it is generally said that glucose cannot be synthesized
3ATPs by oxidative phosphorylation, this is equivalent to from fat. This is because fatty acids are oxidized to acetyl
the input of another six ATPs per glucose synthesized. CoA via -oxidation (Chapter 11). Acetyl CoA cannot be
In spite of being expensive, gluconeogenesis is fairly converted to pyruvate, because the pyruvate dehydrogenase
efficient—the liver can make a kilogram of glucose per reaction is irreversible and there are no alternative reac-
day by gluconeogenesis. tions to channel acetyl CoA into gluconeogenesis.
However, odd chain fatty acids are exceptions because their
E. Substrates for Gluconeogenesis terminal three carbons may serve as gluconeogenic substrate.
The substances that are degraded to acetyl CoA,
Pyruvate and TCA cycle intermediates are important sub- including ketone bodies, ethanol and even chain fatty
strates. In addition, a variety of other molecules can also acids, are also not substrates of gluconeogenesis.
Table 9.4 Energy expenditure for synthesis of one glucose molecule from two molecules of pyruvate
Enzyme Reaction Energy consumption
Pyruvate carboxylase 2 × Pyruvate 2 × Oxaloacetate 2 ATPs
PEP-carboxy-kinase 2 × Oxaloacetate 2 × PEP 2 GTPs
Phosphoglycerate kinase 2 × 3 Phosphoglycerate 2 × 1,3 Bisphoglycerate 2 ATP
Sum: 6 High energy phosphate bonds
F. Regulation inhibited by ATP, these two pathways are regulated in

a reciprocal manner. For example, when energy charge
Certain bypass steps of gluconeogenesis serve as important of the cell is low, glycolysis is favoured but gluconeo-
control points. genesis is inhibited.
Regulation of Pyruvate to Oxaloacetate Hormonal Regulation

Conversion The above mechanisms bring about short-term regulation
Pyruvate carboxylase, the enzyme catalyzing this bypass of gluconeogenesis. The day-to-day regulation is effected
step is a regulatory enzyme; it is allosterically inhibited by the action of hormones on the amount of the key
by ADP and stimulated by acetyl CoA. So potent is the enzymes. Insulin represses the gluconeogenic enzymes
effect of acetyl CoA that the enzyme is virtually inactive in (and induces most of the glycolytic enzymes); and these
its absence. Stimulation by acetyl CoA ensures that suffi- effects are counter-balanced by glucagon and other insu-
cient oxaloacetate is generated (by pyruvate carboxylase) lin antagonists.
for condensing with it (acetyl CoA) to form citrate
(Fig. 9.25a). Regulation by Glucagon
Glucagon, a peptide hormone produced by the -cells in
Regulation of Fructose 1,6-bisphosphate to the endocrine pancreas stimulates gluconeogenesis by
Fructose 6-phosphate Conversion following mechanisms:
ATP is a positive modulator, while AMP affects fructose
1. Glucagon raises the intracellular concentration of the
1,6-bisphosphatase negatively (Fig. 9.25b). Therefore, rate
second messenger cyclic AMP (cAMP), which causes
of gluconeogenesis is determined by cellular energy
conversion of the active form of pyruvate kinase to the
charge, reflected by the ratio of ATP : (ADP AMP)
inactive form (Fig. 9.9). This reduces the PEP to pyru-
1. When the energy charge is low, as indicated by increased vate conversion and the former is diverted into the
intracellular AMP concentration, rate of gluconeo- gluconeogenic sequence.
genesis is impeded through inhibition of fructose 2. Glucagon reduces the intracellular concentration of
1,6-bisphosphatase by AMP. fructose 2,6-bisphosphate (Fig. 9.8). This compound
2. Conversely, when the energy charge is high, rate of is a negative allosteric modulator of fructose 1,6-
gluconeogenesis is enhanced due to stimulation of bisphosphatase and, as explained earlier, a positive
this enzyme by ATP. Since the corresponding glycolytic modulator of phosphofructokinase (PFK1); both these
enzyme, PFK1, is stimulated by AMP and ADP, and factors ultimately favour gluconeogenesis.
Glucose
(a) (b)
Glucose-6-P
Pyruvate
− AMP
Fructose-6-P
PDH − Fructose-
PC bisphosphatase Fr-2,6-BP
ADP − Fructose 1,6-bis-P

+ + ATP
Acetyl CoA Common steps
Oxaloacetate ADP
Phosphoenol pyruvate Citrate, H+
inhibits
Pyruvate
PEP carboxy PC
kinase
Oxaloacetate
TCA
cycle
Fig. 9.25. Regulation of gluconeogenesis. (a) Acetyl CoA as the positive allosteric modulator of pyruvate carboxylase (PC) and
inhibitor of pyruvate dehydrogenase (PDH) complex, (b) Role of various other allosteric modulators ( − inhibits reaction
+ accelerates reaction, Fr-2,6-BP fructose 2,6-bisphosphate).
3. Glucagon increases fatty acid mobilization and their is inhibited in fed state. During starvation, glyco-
oxidation to yield acetyl CoA, NADH and ATP; these neogenesis predominates because levels of fructose
compounds inhibit the PDH complex and acetyl CoA 2,6-bisphosphate are low (refer to Box 9.7 for details).
stimulates pyruvate carboxylase (Fig. 9.25a). Thus, the
available pyruvate is not converted to acetyl CoA but GN Ratio or DN Ratio
to oxaloacetate for gluconeogenesis. It refers to the ratio between the dextrose (glucose) to nitro-
4. Glucagon causes induction of the key enzymes of glu- gen excreted in urine. In the experimental animals treated
coneogenesis, i.e. phosphoenolpyruvate carboxykinase and with alloxan (causes destruction of pancreatic -cells) or
glucose 6-phosphatase (and possibly pyruvate carboxyl- phlorizin (causes renal tubular cell necrosis) glucose is
ase also).
Glycolysis Gluconeogenesis
Regulation by Insulin Fr-6-phosphate
Insulin causes repression of severed gluconeogenic enzymes,
a detailed account of which is given in Chapter 15. Citrate − + Citrate
AMP + − Fr-2,6-BP
Reciprocal Regulation of Gluconeogenesis and
Fr-2,6-BP + − AMP
Glycolysis ATP −
It is evident from the above discussion that glucagon not
only stimulates gluconeogenesis but concomitantly inhib- Fr-1,6-bisphosphate
its glycolysis. Allosteric modulations also ensure that the
opposing enzymes of these two processes are not active
at the same time, which in turn prevents futile cycles
(refer to Box 9.8 for details):
1. As shown in Figure 9.26, compounds reflecting high PEP

energy state of cell, e.g. ATP, citrate and acetyl CoA, allo- ATP − − ADP
sterically stimulate the key gluconeogenic enzymes
but inhibit the corresponding enzymes of glycolysis. OAA
2. Fructose 2,6-bisphosphate also has opposite effects − ADP
on phosphofructokinase (stimulatory) and fructose + Acetyl CoA
Pyruvate
1,6-bisphosphatase (inhibitory). Levels of fructose 2,6-
bisphosphate are high in fed state (and low in starvation) Fig. 9.26. Reciprocal regulation of glycolysis and gluconeo-
so that glycolysis is accelerated and glyconeogenesis genesis.
BOX 9.7
Gluconeogenesis in Starvation and Well-fed State
Intramitochondrial concentration of acetyl CoA rises in starvation due to suppression of TCA cycle and stimulation of
-oxidation (Chapter 15). Acetyl CoA causes stimulation of the pyruvate carboxylase, which accelerates the pyruvate to
oxaloacetate conversion. Since acetyl CoA inhibits activity of pyruvate dehydrogenase, pyruvate to acetyl CoA conversion
is concomitantly inhibited. Thus, most of the available pyruvate is directed to oxaloacetate formation (Fig. 9.25).
Oxaloacetate, so formed, is mostly channeled into gluconeogenic sequence for the generation of glucose (OAA cannot
enter the TCA cycle since this pathway is suppressed in starvation). Generation of glucose in this manner ensures that nor-
mal blood glucose levels are maintained in starvation.
Conversely, when intracellular level of acetyl CoA falls, pyruvate carboxylase becomes inactive. Such situation arises in
the well-fed state, when insulin release is enhanced, which funnels pyruvate into the other available route, i.e. oxidation by
pyruvate dehydrogenase pathway to acetyl (CoA).
In short, during starvation pyruvate is mostly used for the production of OAA, a gluconeogenic substrate, whereas in
well-fed state it is converted to acetyl CoA, which enters TCA cycle and lipogenic sequence.
lost through urine and body attempts to generate glucose Glycogen is an extensively branched homopolysac-
through gluconeogenesis. The body proteins are degraded charide, consisting of chains of (14) linked glucosyl
to provide amino acid substrates for gluconeogenesis, residues with branches that are joined by (16) linkages
and their catabolic end-product, urea, is excreted in urine. (Chapter 2). The latter are spaced about every 4–6 residues
The ratio of glucose to urea nitrogen in such animals is along the -1,4 chain. The gross structure of glycogen is
found to be 3.65, meaning that one gram of nitrogen (from dendritic in appearance, expanding from a core sequence
proteins) forms 3.65 grams of glucose. bound to a tyrosine residue in the protein, glycogenin.
GN ratio is increased when catabolism is enhanced, such
as in starvation, hyperthyroidism and cancer. A. Glycogen Synthesis (Glycogenesis)
Because 3.65 grams glucose is synthesized from one gram
of nitrogen of protein, and proteins contain 16% nitro- Glycogen is synthesized from glucose. Like most other ana-
gen, it is calculated that 58% of protein is glycogenic. bolic processes, glycogenesis occurs in the cytosolic fraction
of the cell and requires input of energy. The energy for gly-
cogenesis comes from high energy nucleotides, ATP and
VI. Glycogen Metabolism UTP. These nucleotides provide energy for the conversion
of the precursor glucose molecule to its energized form,
Glycogen is stored mainly in liver and skeletal muscles as UDP-glucose. The UDP-glucose (UDPG) serves as activated
an energy reserve, just as starch in plants. The tissue con- donor of glucose residues that are added to the growing glyco-
centration of glycogen in liver (6–8%) is higher than that gen molecule. Glycogen synthase is the major enzyme of this
in muscle (1–2%), but because of the relative masses of complex process, which occurs in following four stages:
muscle and liver, the majority of glycogen in the body is
stored in muscles. Though total hepatic glycogen stores Synthesis of UDP-glucose
(50–100 g) are significantly less than the muscle stores Activation of glucose to form UDP-glucose (UDPG) occurs
(200–300 g), they are involved in a vital activity, that is to in three sequential reactions, outlined in Figure 9.27.
serve as our first line of defense against declining blood
glucose level, particularly between meals. Muscle glycogen
Reaction 1
Glucose is phosphorylated at C-6 by glucokinase in liver
is essential for muscle energy metabolism during bursts
(hexokinase in muscle) to form glucose 6-phosphate.
of physical activity even though muscle relies primarily
Glucokinase has high Km (i.e. low affinity for glucose)
on fats as a source for energy.
and high Vmax which permits it to rapidly phosphorylate
large quantities of glucose after meals, when the ambient
Glycogen is the storage form of carbohydrates in liver glucose concentration is high.
and muscles. Hepatic glycogen generates glucose (due
Reaction 2
to presence of glucose 6-phosphatase), whereas muscle
The phosphate group of glucose 6-phosphate is then shifted
glycogen serves as a source of metabolic fuel for use in
from the sixth carbon to the first carbon of the molecule
muscle.
by phosphoglucomutase to form glucose 1-phosphate.
CH2OH ATP ADP H2C O P

O O
HO OH OH Hexokinase OH
HO OH
Glucokinase
OH OH
Glucose Glucose 6-phosphate
Phosphoglucomutase
2Pi
CH2OH
O CH2OH
Uracil PPi UTP O
HO OH
O P P Ribose OH
HO O P
OH UDP-glucose OH
pyrophosphorylase
UDP-glucose
Glucose 1-phosphate
Fig. 9.27. Synthesis of UDP-glucose, the activated form of glucose for glycogen synthesis.
CH2OH CH2OH
CH2OH O O
O
Uracil +
HO OH OH
HO OH O OH
O P P Ribose
OH OH
OH n
Glycogen
UDP-glucose
Glycogen synthase
CH2OH CH2OH CH2OH

O O O
Uracil
OH OH OH
HO O O OH + P P Ribose
OH OH OH
Glycogen (elongated by one glucose residue) n UDP
Fig. 9.28. The glycogen synthase reaction; UDPG serving as an activated donor of a glucose residue.
Reaction 3 Elongation of Chains by Glycogen Synthase

Activation of glucose 1-phosphate to the sugar nucleo- The primer is elongated by sequential addition of
tide, uridine diphosphate glucose (UDPG) occurs by the glucose residues. The enzyme glycogen synthase catalyzes
enzyme UDPG pyrophosphorylase. this reaction. It transfers the glucose residues from UDPG
to the non-reducing end of glycogen in (14) linkage
Glucose 1-phosphate UTP UDPG PPi
(Fig. 9.28).
Pyrophophosphate produced in this reaction is hydro- UDPG Glycogen UDP Glycogen (elongated by
lyzed to inorganic phosphates by pyrophosphatase; this one glucose residues)
ensures irreversibility of the reaction.
This reaction has been elaborately shown in Figure 9.28.
UDP produced in the above reaction is reconverted to
Synthesis of Primer to Initiate Glycogen UTP by the enzyme nucleoside diphosphate kinase.
Synthesis
UDP ATP UTP ADP
UDPG donates glucose residues to an existing (14)
glucosyl chain called primer, which will accept the
incoming glucose residues.
Formation of Branches in Glycogen
The reactions discussed so far result in formation of lin-
UDPG Glycogen primer UDP Glycogen ear, unbranched chains that are linked by (14) link-
(n residues) (n 1 residues) age. However, glycogen is a highly branched structure; the
branch points are created by the action of the “branching
Normally a fragment of glycogen serves as a primer. Such enzyme“. This enzyme comes into action after glycogen
fragments are obtained from glycogen molecules that synthase has added at least ten glucosyl units (Fig. 9.29).
had been partially degraded in liver during fasting or in The branching enzyme removes a block of five to eight
muscle during exercise. However, when the glycogen glucose residues from the non-reducing end of the chain
stores are depleted, a specific protein, known as glyco- and transfers it to a more internal location on the same
genin, provides the site at which the primer is built. or other chain. The transferred block is attached via
Glycogenine contains a specific tyrosyl residue, which (16) linkage to a glucose residue at the new location.
accepts the first glucose donated by UDPG. The branching enzyme is also referred to as a glucosyl
–Tyrosyl–OH UDPG Tyrosyl–O–glucose UDP 4 : 6 transferase because it forms a new (16) bond
instead of an (14) bond.
The above reaction is catalyzed by glycogenin (autoca- The transfer of a block results in creation of a new
talysis) or by the enzyme glycogen synthase. non-reducing end. More of glucose residues can be
More of glucosyl residues coming from UDP glucose, added to this end. In this way, branching increases the
are attached sequentially in the same manner. Thus a number of non-reducing ends to which new glucose
short (14) glucosyl chain (i.e. the primer) is built which residues can be added, thereby greatly accelerating the
is attached to the protein. rate at which glycogen synthesis occurs.
Non-reducing
end
Glycogen Branching
synthase enzyme
New non-reducing
Re end of the transferred
du block
cin
ge
nd
Fig. 9.29. The action of branching enzyme. It forms branches by transferring a block of glycosyl residues, typically seven in
length, from the end of an unbranched chain to a more interior location (on the same or other chain). The transferred block is
attached via 1–6 linkage. indicates non-reducing ends [each circle represents a glucose residue].
(a) (b) (c) (d)
α-1,6 bond α(1→6)-

Glucosidase
Phosphorylase 4 : 4 Transferase
Glucose Glucose
1-phosphate
Reducing
end
Fig. 9.30. Glycogen degradation. Designation of glucose residues as in Figure 9.28.
B. Glycogen Degradation external glucose residues until the branches are about
four residues long. Then the action of glycogen phosphory-
(Glycogenolysis) lase stops and debranching enzyme comes into play.
The principal enzyme of glycogenolysis is glycogen phos-
phorylase. It acts in association with a debranching enzyme
Removal of Branches
The key enzyme for removing branch points of glycogen
(Fig. 9.30).
is the debranching enzyme, which possesses dual activ-
ity, namely glucosyl 4 : 4 transferase activity and a(14)
Action of Glycogen Phosphorylase glucosidase activity.
Glycogen phosphorylase removes glucose residues, one at a
time, from the non-reducing ends of glycogen (Fig. 9.30a). 4 : 4 transferase activity: Three of the four external glucosyl
It utilizes inorganic phosphate (P1) to cleave the (14) residues that remain are removed as a trisaccharide and
bonds. This results in release of the terminal glucosyl res- transferred to the non-reducing end of a nearby chain (Fig.
idue as glucose 1-phosphate. The reaction is termed as 9.30c). This action involves cleaving of an (14) bond at
phosphorolysis, that is, breakage of a covalent bond by one site and formation of a new (14) bond elsewhere.
addition of a phosphate group. ␣(16) glucosidase activity: The single glucosyl residue
that remains at the branch point is removed by the
Glycogen Pi Glycogen Glucose 1-phosphate
(16) glucosidase, to liberate free glucose (Fig. 9.30d).
(n residues) (n1 residues)
Removal of branch point in this manner exposes another
Pyridoxal phosphate is an essential cofactor in this reac- set of (14) linkages, till the next branch point. When
tion; it is covalently bound to the enzyme protein. another branch point is reached, it is again removed by
Phosphorylase is specific for (14) linkages only, it can- the debranching enzyme, and the cycle thus continues.
not cleave (16) linkages. Further, this enzyme cannot Thus, the phosphorolysis/debranching processes, act-
approach the branching glucose residues efficiently. ing alternately, can cleave a large glycogen molecule hav-
Thus, as shown in Figure 9.30b, phosphorylase cleaves the ing thousands of glycosyl residues.
Fate of Glucosyl Units Released from Glycogen Covalent Modulation

The glucosyl units are released from glycogen in two Studies concerning regulation of glycogen phosphorylase have
forms: glucose 1-phosphate and free glucose. About 90% been mostly conducted in skeletal muscles. The muscle
of the glucose is released as glucose 1-phosphate, and only the enzyme is a dimeric protein consisting of two identical
-1,6 branching residue, is released as free glucose. Glucose subunits. Each subunit contains an essential serine resi-
1-phosphate is then converted to glucose 6-phosphate due, where phosphate group can covalently attach. Thus,
by the enzyme phosphoglucomutase. the glycogen phosphorylase exists in two forms:
Further, fate of glucose 6-phosphate varies depending on
Phosphorylated form called phosphorylase a which is
the tissue. Liver possesses the enzyme glucose 6-phosphatase,
catalytically active.
which forms free glucose from glucose 6-phosphate. This
Dephosphorylated form called phosphorylase b which is
glucose is released into blood for use by needy tissues.
much less active.
Muscles lack glucose 6-phosphatase, hence cannot con-
tribute to the blood glucose. Rather muscle glycogen is These two forms of the enzyme are interconvertible.
used for the generation of metabolic energy in the exer- Conversion of inactive phosphorylase b to active phosphorylase a
cising muscles themselves. is a catalyzed by the enzyme phosphorylase kinase, which
Free glucose
phosphorylates, and thereby activates glycogen phosphory-
lase. Another enzyme, protein phosphatase-1, dephos-
Glycogen
phorylates, and thereby inactivates glycogen phosphorylase
Glucose 1-phosphate
a (Fig. 9.31).
E1 Through action of these enzymes, ratio of the active
Glucose 6-phosphate Glucose
and the inactive phosphorylase can be varied, which ulti-
E1 Glucose 6-phosphatase in liver mately controls the rate of glycogenolysis. The same
mechanism is operative in liver as well.
Role of liver glycogen and muscle glycogen are different: Liver
releases free glucose in blood circulation, and thereby plays Allosteric Regulation
an important role in glucose homeostasis. However, hepatic Superimposed upon the covalent modulation are the
glycogen stores are barely sufficient for maintenance of actions of allosteric effectors:
blood glucose concentration during a 12-hour fast.
1. AMP: Glycogen phosphorylase in muscles and other
Muscle glycogen stores are used for liberating fuel mol-
extrahepatic tissues is stimulated powerfully by AMP
ecules (i.e. glucose 6-phosphate), which in turn are used
(Fig. 9.31). This allosteric effect ensures that glycogen
in the exercising muscles for generating metabolic energy.
is degraded rapidly in severely contracting muscles,
to provide substrate for an anaerobic glycolysis.
C. Regulation Concentration of AMP quickly builds up in contract-
ing muscles due to its rapid production by pyrophos-
Synthesis and breakdown of glycogen are important phate cleavage of ATP
intracellular activities, having implications beyond the ATP AMP PPi
cell. Balance between the two processes is important for:
Note: Unlike other forms of stored energy, only gly-
(a) sustaining adequate glycogen stores, and cogen can be used for ATP synthesis under anaerobic
(b) for maintaining the normal blood glucose levels. conditions.
Normal blood glucose levels are especially important 2. Glucose: Glycogen phosphorylase in liver is inhibited by
for the tissues that use glucose as their primary substrate, glucose. Because the intracellular glucose concentra-
such as, brain erythrocytes, renal medulla, lens and cor- tion in the liver approximates the blood glucose level,
nea of the eye. Therefore, tight regulation of these two pro- glycogen degradation in liver is regulated directly by
cesses (i.e. glycogenesis and glycogenolysis) is essential. the blood glucose level.
3. Glucose 6-phosphate: It inhibits muscle glycogenolysis.
4. ATP: Both liver and muscle glycogenolysis are inhib-
Regulation of Activity of Glycogen
ited by ATP.
Phosphorylase
The regulatory enzyme for glycogenolysis is glycogen phos- Glucose Glycogen phosphorylase (Liver)
−
phorylase. Activity of this enzyme is regulated by (a) covalent
AMP Glycogen phosphorylase (Muscle)
modulation through phosphorylation-dephosphorylation, +
(b) allosteric regulation modulation, and (c) calcium ions. Glucose 6-phosphate −
AMP Glucose 6- P ATP ADP Glucose

(liver)
Phosphorylase kinase
+ − −
Glycogen phosphorylase b Glycogen phosphorylase a

(Less active) (Fully active)
Protein phosphatase-1 Phosphorylated enzyme

Dephosphorylated enzyme
Pi
Fig. 9.31. Regulation of glycogen phosphorylase by covalent modification and allosteric effectors.
(Fig. 9.32). Thus, it is activated by protein kinase and ren-

Table 9.5 Difference between glycogenolysis in liver and
muscles dered less active by protein phosphatase-1.
It is important to note that simultaneous phosphorylation
Muscle Liver
of both glycogen-phosphorylase and synthase will switch the
1. Little glucose formed, since Free glucose forms by
cell from glycogen synthesis to glycogen degradation. This pre-
glucose 6-phosphatase is glucose 6-phosphatase
vents futile cycles (Box 9.8).
absent
2. Stimulated by adrenaline, Glucagon is major stimulator
glucagon has no effect of glycogenolysis Allosteric Regulation
3. Stimulated by AMP but No effect of AMP, but hepatic Glucose 6-phosphate is a potent activator of glycogene-
inhibited by glucose glycogen phosphorylase is sis. It does so by stimulating the protein phosphatase-1, so
6-phosphate inhibited by glucose that glycogen synthase is converted to the active (dephos-
phorylated) form. It also causes direct allosteric stimula-
To sum up, the various mechanisms described ensure tion of the glycogen synthase activity. In muscle tissue,
that glycogenolysis is stimulated when glucose concentration glucose 6-phosphate not only stimulates glycogen syn-
and energy levels are low, and inhibited when these are high. thase, but also inhibits activity of glycogen phosphorylase.
Regulation by Calcium Ions

During muscle contraction, calcium is released from sar- Availability of glucose 6-phosphate is high in fed state,
coplasmic reticulum and activates phosphorylase kinase which ensures activation of glycogenesis and hence gly-
(via a calmodulin-calcium modulating protein; Chapter
cogen deposition.
29). This enzyme causes phosphorylation of glycogen phos-
phorylase and hence enhances glycogenolysis (without
involving cAMP). Hormonal Regulation of Glycogen Metabolism
Hepatic and muscle glycogenolysis serve different Glucagon and epinephrine, acting through cAMP, are pri-
roles and respond to different regulatory signals. marily involved in regulation of the glycogen metabolism.
Major differences between the two are given in Table 9.5. Glucagon acts on liver cells and epinephrine acts on both
liver and muscle cells. These hormones stimulate glycoge-
Regulation of Activity of Glycogen Synthase nolysis and inhibit glycogenesis. Insulin, on the other hand,
The regulatory enzyme of glycogenesis is glycogen synthase. promotes glycogenesis and inhibits glycogenolysis.
Regulation of its activity is effected by the following
mechanisms: (a) allosteric regulation, and (b) covalent Role of Glucagon and Epinephrine
modulation. Both these hormones activate the membrane enzyme
adenylate cyclase, which catalyzes formation of cAMP from
Covalent Modulation ATP (Fig. 9.33). The cAMP binds to protein kinase A (PKA,
Activity of glycogen synthase is also regulated by also called cAMP-dependent protein kinase), a tetrameric
phosphorylation-dephosphorylation, as in the case of protein. The cAMP binding activates this enzyme by let-
glycogen phosphorylase. However, there is a major differ- ting its catalytic subunits free (Chapter 29). Activation
ence: glycogen synthase is more active in the dephosphory- of PKA has the following consequences: (i) Inhibition of
lated form and less active in the phosphorylated form glycogenesis, and (ii) Stimulation of glycogenolysis.
ATP ADP
Protein kinase Glucose 6- P
+
Glycogen synthase a Glycogen synthase b
(Fully active) (Less active)
Protein phosphatase-1
Dephosphorylated enzyme Phosphorylated enzyme
Pi
Fig. 9.32. Regulation of glycogen synthase by covalent modification and allosteric effectors.
BOX 9.8
Futile Cycles
The synthetic and the degradative pathways of metabolism are fine-tuned by allosteric effectors and hormone-induced
enzyme phosphorylation in order to meet the cellular demands for metabolites and energy, and at the same time minimize
their wastage. Normally, opposing pathways (e.g. glycolysis and gluconeogenesis; glycogen synthesis and its degradation)
are reciprocally regulated: activation of one is accompanied by inactivation of the other, and vice versa. This is necessary
because simultaneous activity of these opposing pathways would lead to a wasteful metabolic exercise, called futile cycle,
that achieves little but ATP hydrolysis.
Futile cycles are minimized through reciprocal regulation of the opposing enzymes. For example, the key enzymes of
glycogen synthesis (glycogen synthase) and glycogen degradation (glycogen phosphorylase) can both be phosphorylated
on specific serine side chains by protein kinases (and dephosphorylated by protein phosphatase). Glycogen synthase is more
active in the dephosphorylated form and glycogen phosphorylase is more active in the phosphorylated form. Therefore,
the simultaneous phosphorylation of both enzymes will switch the cell from glycogen synthesis to glycogen degradation.
Conversely, simultaneous dephosphorylation will switch the cell from glycogen degradation to glycogen synthesis, simulta-
neous activation of both (i.e. futile cycling) being prevented in either case.
Inhibition of glycogenesis: The active PKA cause phos- in activated (phosphorylated) form providing a
phorylation of glycogen synthase. Since the phosphor- persistent stimulus for glycogenolysis (Fig. 9.34).
ylated glycogen synthase is less active, glycogenesis is
inhibited. Note: Inhibition of protein phosphatase-1 locks glycogen
Stimulation of glycogenolysis: The active PKA stimulates synthase in its inactive (phosphorylated) form, hence
glycogenolysis by a dual mechanism, as discussed below: suppressing glycogenesis.
(a) It catalyzes phosphorylation of phosphorylase
kinase, which exists as an inactive dephosphory- Cascade with amplification: The cAMP activated process
lated form and an active phosphorylated form. is a cascade in which the initial hormonal signal is ampli-
Thus, phosphorylase kinase is now activated, and in fied several folds. Single hormone molecule of glucagon
turn phosphorylates the glycogen phosphorylase b, or adrenaline activates the enzyme adenylate cyclase for
converting it to phosphorylase a, that now carries long enough to produce hundreds of cAMPs which acti-
out a rapid dehydration of glycogen (Fig. 9.33). vate protein kinase molecules. Although four cAMP mol-
(b) Active PKA phosphorylates a protein called ecules are required to activate a protein kinase (Chapter
inhibitor-1, which thereby gets stimulated. This pro- 29), each active PKR molecule can convert hundreds or
tein inhibits protein phosphatase-1, the enzyme that thousands of molecules of phosphorylase kinase to active
normally causes inactivation of glycogen phosphor- form. Similarly, each active molecule of phosphorylase
ylase a. The overall effect is that the glycogen phos- kinase, can convert thousands of molecules of glycogen
phorylase a dose not get inactivated, and is “locked” phosphorylase b to phosphorylase a, and so on. The net result
Plasma
Receptor
membrane
ATP cAMP + PPi

Adenylate cyclase
Protein kinase (inactive) Protein kinase (active)
Glycogen Glycogen
synthase a synthase b
(active) (inactive)
+ Glycogenesis
inhibited
Phosphorylase
+ kinase (inactive) Phosphorylase
kinase (active)
+
Protein
phosphatase-1
Glycogen Glycogen
phosphorylase b phosphorylase a
− (inactive) (active)
Inhibitor-1
Protein Glycogenolysis
− phosphatase-1 stimulated
Fig. 9.33. Hormonal regulation of glycogen synthesis and degradation. The cAMP generated by hormones activates protein
kinase A, which cause phosphorylation of glycogen synthase (inactivation), phosphorylase kinase (activation) and the inhibitor-1
(activation). The last inhibits protein phosphatase. Activated phosphorylase kinase causes phosphorylation of glycogen phosphorylase b,
activating it to glycogen phosphorylase ( inhibitor, stimulator).
+ − Protein phosphatase. The latter causes dephosphorylation of gly-

Protein kinase A Inhibitor-1 Phosphatase-1
cogen synthase. Dephosphorylation causes this enzyme to
+ be activated, resulting in increased glycogen synthesis.
Phosphorylase Inactivates Glycogen Glycogen Moreover, insulin stimulates the uptake of glucose by
kinase glycogen phosphorylase synthase locked muscle, providing substrate molecules for glycogen syn-
synthase locked in in inactive form
activated form thesis. Dephosphorylation of glycogen phosphorylase causes
Activates glycogen a concomitant fall in the rate of glycogenolysis. Finally,
phosphorylase insulin promotes activity of phosphodiesterase (it degrades
Fig. 9.34. Dual control of glycogenesis and glycogenolysis by cAMP) in liver, which decreases cAMP levels. This
protein kinase A. enhances glycogenesis and suppresses glycogenolysis.
is that a single hormone molecule can generate thousands of

Glycogen is synthesized in pathway of glycogenesis (major
molecules of glucose 1-phosphate by glycogenolysis.
enzyme, glycogen synthase) and degraded by phospho-
rolytic cleavage (major enzyme, glycogen phosphorylase)
Role of Insulin
in pathway of glycogenolysis. The two pathways are
Insulin is the key hormone that stimulates glycogen synthe-
reciprocally regulated by hormones and metabolites.
sis. It is released in the fed state and activates an intracellular
D. Glycogen Storage Diseases Table 9.6. Glycogen storage diseases

Type Enzyme deficiency Organ (s)
Any deficiency of a glycogen degrading enzyme causes affected
abnormal accumulation of glycogen in the affected tissue. I von Gierke’s Glucose 6-phosphatase Liver, kidney
The resulting diseases, summarized in Table 9.6, are col- disease
lectively known as the glycogen storage diseases. These II Pompe’s (14) Glucosidase All organs
diseases are quite rare (overall incidence of 1 in 40 000) disease (acid maltase)
and are mostly inherited as autosomal recessive trait.
III Cori’s disease Debranching enzyme Muscle, liver
Normally a defect in an enzyme of glycogenolysis (or
IV Andersen’s Branching enzyme Liver,
glycogenesis) leads to abnormal accumulation of glyco-
disease myocardium
gen. However, defective action of a glycolytic enzyme,
V McArdle’s Phosphorylase Muscle
phosphofructokinase, may also result in abnormal glyco-
disease
gen accumulation.
VI Hers’ disease Phosphorylase Liver
In some forms, the main abnormality lies in liver,
VII Tarui’s disease Phosphofructokinase Muscle, RBCs
whereas in others muscle tissue is primarily affected.
VIII Phosphorylase kinase* Liver
Accordingly, glycogen storage diseases can be classified
* There is also on X-linked form of phosphorylase kinase deficiency. This is
in two major types: the hepatic forms and the myopathic
sole exception as all other glycogen storage diseases are inherited as
forms. Certain forms (e.g. Type II) cannot be put into autosomal recessive trait.
either of these categories. Some rare forms, e.g. IX, X and
XI have also been identified which are due to defects in
2. Hyperuricaemia: It may develop because accumula-
enzymes that activate or deactivate liver phosphorylase.
tion of glucose 6-phosphate enhances the reactions of
HMP shunt (2nd stage). This stage involves interconver-
Hepatic Forms sion of hexose phosphates (e.g. glucose 6-phosphate)
Type I, III, IV, VI and VII are classified as the hepatic forms and pentose phosphates (e.g. ribose 5-phosphate).
of glycogen storage disease. These are characterized by Therefore, accumulation of glucose 6-phosphate leads
excessive deposition of glycogen in liver, and hence liver to increased formation of ribose 5-phosphate. This
enlargement (hepatomegaly) is the prominent feature. The leads to increased uric acid production since ribose
biochemical manifestations are hypoglycaemia, lactic aci- phosphate enhances formation of purine nucleotides
dosis, hyperuricaemia and hyperlipidaemia, which are most by “salvage pathway” (Chapter 20), which are then
prominent in the type I form (von Gierke’s disease), but degraded to uric acid.
may occur in varying severities in the other forms. Enzy- Moreover, lactic acidosis inhibits renal excretion of
matic defects in various forms are shown in Table 9.6. uric acid, which aggravates hyperuricaemia.
von Gierke’s disease (Type I): It is a classical hepatic gly-

Glucose 6-phosphate Glucose
cogen storage disease, caused by deficiency of glucose accumulation
6-phosphatase. It is a rare disorder with an incidence of Glycolysis Pyruvate ↑Lactate
HMP
1 per 200,000 persons. It is characterized by severe hepa- shunt
tomegaly and dangerous hypoglycaemia, that may develop ↑Pentose phosphates Purines ↑Uric acid
Salvage
within hours of last meal (not surprising, because in pathway
between meals glucose 6-phosphatase is required for for-
mation of glucose by both glycogenolysis and gluconeo- 3. Hyperlipidaemia: It is the result of decreased glucose
genesis). The patient may be kept alive only by feeding availability, leading to mobilization of the stored fats.
carbohydrates at regular intervals, day and night. The hypogly-
The above features may occur, in varying degrees, in the
caemia is unresponsive to glucagon or adrenaline because
other hepatic forms of glycogen storage diseases (Table 9.6).
of the metabolic block in formation of free glucose from
liver glycogen stores. Other manifestations of von Cori’s disease (Type III): Branched chain glycogen accu-
Gierke’s disease are as follows: mulates because of absence of the debranching enzyme.
Clinical course runs like the type I, but is much milder.
1. Lactic acidosis: It may develop because the glucose
6-phosphate cannot be degraded to glucose, and is Anderson’s disease (Type IV): A rare disease in which gly-
therefore, diverted into the glycolytic sequence to cogen having few branches accumulates because of
form pyruvate and then lactate. Lactate level in blood absence of the branching enzyme. Death from liver cir-
increases and pH is lowered (acidosis). rhosis usually occurs before the age of two years.
Her’s disease (Type VI): Liver glycogen stores cannot be Exercises

degraded due to phosphorylase deficiency, so liver enlarges
and mild hypoglycaemia and ketosis are often seen. The Essay type questions
disease runs a mild course. 1. Enumerate the rate regulating enzymes of glycolysis
and glyconeogenesis. How are these enzymes regu-
Type VIII: Mild hepatomegaly and hypoglycaemia char-
lated in a reciprocal manner?
acterize this condition.
2. Describe glycolysis and its regulation. Explain ener-
getics of aerobic and anaerobic glycolysis.
Myopathic Forms 3. List distinctive features of glucokinase and hexokinase.
These include types V (McArdle’s disease) and VII (Tauri’s
Explain why the high Km of glycokinase is important
disease). Muscle glycogen stores are very high but are not
for the role of the liver in buffering blood glucose.
available for use. Because glycogen is the primary sub-
4. Describe the conditions under which the Cori’s cycle
strate in the exercising muscles, the latter are most severely
and the glucose-alanine cycle operate.
affected. Clinical course of both type V and type VII runs
5. Describe the reactions of the pyruvate dehydrogenase
similarly: the affected patients suffer from muscle cramps
multienzyme complex. How is entry of acetyl CoA
and pain upon exertion and are easily fatigued. Apart
into the TCA cycle regulated at this complex?
from this they lead a normal life. This shows that the
6. Mention the reactions of glycogenesis and glycogenol-
utilization of muscle glycogen is not essential for life.
ysis in flow diagram. Describe how these pathways
Interestingly, after muscular activity, these patients do
are regulated. Describe the distinctive features of gly-
not show the expected increase in blood lactate. This
cogen synthesis and degradation in liver and skeletal
demonstrates that the most important source of lactic
muscle.
acid during muscular activity is not the free glucose from
7. With at least one example each related to glucose
blood but stored muscle glycogen.
metabolism, discuss how enzyme activity is regu-
Pompe’s disease (Type II), which involves all organs, lated by (a) allosteric mechanism, and (b) covalent
needs a special mention because of its peculiar features. In modification.
this disease, glycogen accumulation occurs in lysosomes 8. Highlight the role played by protein kinases in glycogen
since ability to hydrolyze it is impaired. This is because metabolism. Explain the differences in glycogenolysis
of impairment (or absence) of the lysosomal enzyme between liver and muscle.
a(14) glucosidase. Consequently the lysosomes get dis-
tended with glycogen, especially in the myocardium, skel-
Write short notes on
1. Cori’s cycle
etal muscle, liver and motor nuclei of spinal cord.
2. Glycogen storage diseases
The infantile form of this disease is very severe, and
3. Anaerobic glycolysis
the child dies in first few months of life due to cardiac
4. Pasteur effect
enlargement, that leads to cardiac failure. The juvenile
5. McArdle’s disease
form of this disease is also fatal, but the adult form is
6. GLUTs
relatively milder, being marked by slowly developing
7. Fluoride as inhibitor of glycolysis
myopathy.
8. Essential fructosuria
9. BPG shunt
10. Congenital galactosaemia
Glycogen accumulates in several rare enzyme deficiencies,
11. Renal glycosuria
collectively termed glycogen storage diseases. These dis-
12. Distinctive features of glucokinase and hexokinase
eases (except one) are inherited as autosomal recessive
13. Metabolism of amino sugars
traits.
14. Glyoxylate pathway
CLINICAL CASES
CASE 9.1 A 48-year-old man in a comatose state
A 48-year-old moderately obese man was brought to the and his speech was incoherent. Soon afterwards, he lost con-
medical emergency in an unconscious state. He was breath- sciousness and was rushed to the nearest hospital.
ing deeply and rapidly. The breath did not smell of ace- Blood and urine samples were obtained for analysis.
tone or alcohol. Signs of mild dehydration, such as dry
tongue, soft eyeballs, weak and rapid pulse and low blood Investigations Patient’s Reference
pressure were present. test reports range
Examination of the past medical record showed that the Venous plasma 85 mg/dL 140 mg/dL
patient was diagnosed as having type 2 diabetes mellitus glucose (random)
(NIDDM) six years back. Initial treatment with sulphonyl- pH 7.32 7.35–7.45
ureas proved effective in controlling blood glucose levels,
Plasma bicarbonate 20 mmol/L 21–28 mmol/L
and the patient apparently remained in good health all
these years. However, about four months back, elevated pCO2 32 mmHg 36–45 mmHg
blood glucose levels were detected on two separate occa- Lactic acid 9.8 mmol/L 0.44–1.4
sions (210 mg/dL and 236 mg/dL). Increasing the dose of mmol/L
sulphonylureas did not help. Biguanide therapy was initi- Urine examination: Normal: sugar, proteins or ketone
ated, after the renal and the hepatic functions were found bodies absent.
normal. Adequate control of diabetes was achieved with
metformin in three daily doses of 1.0 g each. Q.1. What is the biochemical abnormality in this patient?
For the last two days, the patient had hectic sessions of Q.2. Diagnose the clinical condition that might have led to
physical activity when he went to a hill station for recreation. the above biochemical abnormality?
He had rich meals during this period, often accompanied by Q.3. What is the role of the following in causing the
drinking till late night. While driving back home early in the patient’s problems: (a) severe muscular exercise, (b)
morning, the family members noticed that he was disoriented acute alcohol intoxication.
CASE 9.2 An unconscious 5-year old child

A 5-year-old child was brought to the medical OPD in a found in the test result. The child was given intravenous
comatose state. He had felt headache and dizziness only a dextrose to treat hypoglycaemia. He responded well and
few hours before. His father noticed profuse sweating and the hypoglycaemic symptoms promptly disappeared. Two
some abnormal behavior at that time and rushed him to the days later, liver biopsy was performed and the sample was
hospital. A similar episode had occurred two months earlier analyzed. It revealed large deposits of fructose 1-phosphate
after the child had a glass of sugarcane juice. However, he within the hepatocytes.
had not lost consciousness on that occasion; his mother had
made him drink a glass of heavily sugared milk thinking Q.1. What is the probable biochemical defect in this
that the child was feeling weak. This seemed to have allevi- child? Suggest a biochemical test to evaluate the
ated the symptoms. above diagnosis.
Presently, physical examination showed tachycardia Q.2. Provide a biochemical explanation for the child’s
and rapid breathing. Liver was markedly enlarged, being signs and symptoms.
palpable 4 cm below costal margin. Blood and urine sam- Q.3. Would you expect liver function to be normal or
ples were obtained for biochemical analysis. Blood glucose sluggish in this child? Give reason.
level was very low (52 mg/dl). No other abnormality was Q.4. Suggest treatment for this condition.
METABOLISM OF
CHAPTER
CARBOHYDRATES II:
SECONDARY PATHWAYS
AND REGULATION OF
BLOOD GLUCOSE LEVEL
10
The mainline metabolic pathways of glucose utilization generate ATP energy and fuel molecules. In addition to such path-
ways, there are other minor (or secondary) pathways taken by glucose, which are specialized for synthesis of glycolipids,
glycoproteins and other special products needed by the cells. Two such pathways are pentose phosphate pathway,
which leads to formation of ribose 5-phosphate and NADPH; and uronic acid pathway, which generates D-glucuronate
and ascorbic acid. Rate of flux of metabolic intermediates through the secondary pathways is much less compared to that
through the mainline pathways.
Significance of certain other minor metabolic pathways that metabolize sugars other than glucose, such as fructose,
galactose, mannose, or the disaccharides and the polysaccharides is also outlined this chapter. Maintenance of blood
glucose level within the normal range has been discussed.
After going through this chapter, the student should be able to understand:
Reactions of pentose phosphate pathway and uronic acid pathway, and their significance.
Minor pathways of metabolism of carbohydrates other than glucose.
Metabolic significance and clinical consequences of enhanced rate of sorbitol pathway in diabetes mellitus.
The interplay of various factors that are responsible for maintenance of normal blood glucose levels.
I. Pentose Phosphate Pathway and cornea of the eye. It accounts for about 60% of the
total oxygen consumption in cornea.
The pentose phosphate pathway also called hexose The pathway consists of three irreversible oxidative
monophosphate (HMP) shunt or phosphogluconate reactions (Stage I), followed by a series of reversible sugar-
pathway is required for provision of five-carbon sugars phosphate interconversions (i.e. non-oxidative reactions
and NADPH. Activity of this cytosolic pathway is minimal Stage II). An overview is presented below.
in muscle and brain, where almost all of the glucose is 1 2 3
Glucose 6-P Pentose phosphates Glycolytic
degraded by glycolysis. However, it is highly active in liver,
intermediates
adipose tissue, adrenal cortex and lactating (but not the non- Stage I Stage II
lactating) mammary gland. These tissues are involved in
synthesis of cholesterol and fatty acids, the processes The Stage I, irreversible oxidative reactions result in
dependent on NADPH. Because NADPH is important for formation pentose phosphates, while a series of reversible
the antioxidant defenses of the body, the pentose phos- non-oxidative interconversions take place in the Stage II,
phate pathway is well developed in cells that are exposed which convert these pentose phosphates into glycolytic
to a high oxygen partial pressure, such as erythrocytes intermediates.
A. Specialized Products Generated by (Fig. 10.1). Two NADPH molecules are also generated
during this reaction sequence comprising of three irre-
Pentose Phosphate Pathway versible oxidative reactions. The reactions are shown in
Figure 10.2.
1. Reduced nicotinamide adenine dinucleotide phosphate
(NADPH) serves as a biochemical reductant in a num-
Dehydrogenation of glucose 6-phosphate
ber of reactions. Such reductive reactions form impor-
Glucose 6-phosphate is irreversibly converted to 6-
tant steps of several biosynthetic pathways, such as
phosphogluconolactone by the enzyme glucose 6-
steroidogenesis and lipogenesis. Therefore, rate of
phosphate dehydrogenase. NADP⫹ as a co-enzyme is specific
the pentose phosphate pathway is high in tissues that
for this reaction. NADP⫹ acts as an acceptor of the reduc-
synthesize steroids and fatty acids.
ing equivalents (i.e. a pair of hydride) that are removed
NADPH generation through HMP shunt is essen-
from the glucose 6-phosphate molecule and gets con-
tial for maintaining integrity of the erythrocyte mem-
verted to NADPH.
branes because glutathione reduction is brought about
This step is the regulatory step of the pathway. NADPH
by NADPH (Case 10.1). These functions are discussed
acts as a negative modulator, causing competitive inhibi-
in detail later in this chapter.
tion of glucose 6-phosphate dehydrogenase (G6PD).
2. Ribose 5-phosphate is an important constituent of the
purine and pyrimidine nucleotides, that perform a vari- Hydrolysis of 6-phosphopgluconolactone
ety of important functions. It is an essential structural 6-Phosphogluconolactone is converted to 6-phospho-
component of various coenzymes such as coenzyme-A, gluconate by addition of a water molecule. This reaction
FAD and NAD⫹. Ribose 5-phosphate can be recon- is catalyzed by the enzyme gluconolactonase and is irre-
verted to hexose phosphates through reversible sugar- versible, like the previous step.
phosphate interconversions by the non-oxidative
reactions of the pentose phosphate pathway. This pro- Formation of ribulose 5-phosphate
vides an important mechanism for the metabolism Oxidative decarboxylation of 6-phosphogluconate, cata-
of five carbon sugars. lyzed by 6-phosphogluconate dehydrogenase, then yields the
ketose sugar, ribulose 5-phosphate, plus a molecule of
NADPH.
B. Reactions of Pentose Phosphate The non-equilibrium nature of the Stage I reactions
Pathway enables the cell to maintain a cytoplasmic ratio of NADPH :
NADP ⴙ ⫽ 100. Interestingly, the ratio of NADH : NAD ⫹
Stage I: Oxidative Phase in the cytoplasm is nearly the inverse, less than 0.01. For
The oxidative reactions of Stage I bring about conver- this reason the cells employ NADPH rather than NADH
sion of glucose 6-phosphate to ribulose 5-phosphate whenever a strong reducing agent is required.
Glycolysis
Glucose 6-phosphate
NADP+ Fructose 6-phosphate
Glyceraldehyde
NADPH + H+ 3-phosphate
(non-oxidative)
(oxidative)
6-phosphogluconolactone
STAGE II
STAGE I
Lipogenesis
Steroidogenesis
Glutathione reduction
Other reactions
6-phosphogluconate
NADP+
Xylulose
NADPH + H+ 5-phosphate
Ribulose 5-phosphate Ribose
5-phosphate
Nucleotide
biosynthesis
Fig. 10.1. Overview of the pentose phosphate pathway.

Metabolism of Carbohydrates II: Secondary Pathways and Regulation of Blood Glucose Level 197
H O O O
C C C O−
NADP+ NADPH + H+ H2O
H C OH H C OH H C OH
HO C H HO C H O HO C H
H C OH H C OH H C OH
Glucose 6-phosphate
Gluconolactonase
H C OH dehydrogenase H C H C OH
2− 2− 2−
CH2OPO3 CH2OPO3 CH2OPO3
Glucose 6-phosphate 6-phosphogluconolactone 6-phosphogluconate
CH2OH NADPH + H+ NADP+

CO2
C O
H C OH
H C OH 6-phosphogluconate
2− dehydrogenase
CH2OPO3
Ribulose 5-phosphate
Fig. 10.2. The oxidative reactions of pentose phosphate pathway.
The ribulose 5-phosphate, the end product of the aldose), formation of a seven-carbon sugar (sedoheptulose
Stage I, is convertible to other pentose phosphates: xylu- 7-phosphate) and a three carbon-sugar (glyceraldehyde
lose 5-phosphate and ribose 5-phosphate. The enzymes 3-phosphate) results.
for these reactions are:
C5 ⫹ C5 C7 ⫹ C3
1. Epimerase, which converts ribulose 5-phosphate to
Thiamine pyrophosphate, a coenzyme for transketolase,
xylulose 5-phosphate.
serves as transient carrier of the two carbon units in this
2. Ketoisomerase, which catalyzes conversion of ribulose
reaction.
5-phosphate to ribose 5-phosphate.
Xylulose Reaction 2
e 5-phosphate
imeras This is catalyzed by transaldolase, which transfers a three-
Ep
carbon unit from the sedoheptulose 7-phosphate to glyc-
Ribulose
5-phosphate
eraldehyde 3-phosphate. This results in the formation of
Keto a erythrose 4-phosphate and fructose 6-phosphate.
isom
eras
e Ribose C7 ⫹ C3 C4 ⫹ C6
5-phosphate
Reaction 3
Stage II: Non-oxidative Interconversion This is again catalyzed by transketolase. The tetrose phos-
Phase phate (erythrose 4-phosphate) generated in the previous
The pentose phosphates generated in Stage I, ribose 5- step, accepts a two-carbon unit from a new pentose
phosphate, xylulose 5-phosphate and ribulose 5-phosphate, phosphate (xylulose 5-phosphate) molecule to form a
are converted to glycolytic intermediates in the second fructose 6-phosphate and glyceraldehyde 3-phosphate.
stage of HMP (Fig. 10.3). This stage comprises a series of
C4 ⫹ C5 C6 ⫹ C3
non-oxidative reversible interconversions, as noted earlier.
The enzymes of this stage are transketolase and transal- Net result of these reactions is formation of two hex-
dose, which catalyze the following three reactions: ose phosphates (fructose 6-phosphate) and a triose
phosphate (glyceraldehyde 3-phosphate), which directly
Reaction 1 enter the glycolytic sequence.
This reaction is catalyzed by transketolase, which transfers Thus, the non-oxidative interconversions (of the
a two carbon unit from a ketose sugar to an aldose sugar. Stage II) permit conversion of the pentoses (generated in
When the two-carbon unit is transferred from a xylulose Stage I) into intermediates of glycolysis. This establishes
5-phosphate (a 5-C ketose) to ribose 5-phosphate (a 5-C a link between the pentoses and the other metabolizable
Xylulose-5-P Ribose-5-P Xylulose-5-P

(C5) (C5) (C5)
Transketolase
Reaction 3
Reaction 1
Glyceraldehyde-3-P Sedoheptulose-7-P
(C3) (C7)
Transaldolase Erythrose-4-P Transketolase

(C4)
Reaction 2 Fructose-6-P
Fructose-6-P Glyceraldehyde-3-P
(C6) (C6)
(C 3)
Glycolysis
Fig. 10.3. The non-oxidative reactions of pentose phosphate pathway. The transketolase transfers a 2-carbon unit, and the trans-
aldolase causes transfer of a 3-carbon unit.
C5 ⫹ C5 C7 ⫹ C3 (Reaction 1; Transketolase)
Sum: C5 ⫹ C5 ⫹ C5 C6 ⫹ C6 ⫹ C3
sugars, thereby integrating the HMP shunt pathway with Activity of glucose 6-phosphate dehydrogenase is competi-
glycolysis. tively inhibited by NADPH, as mentioned earlier. It is the
A summary of reactions of the pentose phosphate path- ratio of NADPH : NADP ⫹ that determines the overall rate
way, as shown in Figures 10.2 and 10.3 can be written as: of the pathway. Under normal circumstances, the ratio of
3 Glucose 6-phosphate ⫹ 6NADP⫹ NADPH/NADP⫹ is sufficiently high (up to 100), which
2 Fructose 6-phosphate ⫹ Glyceraldehyde 3-phosphate ⫹ keeps the activity of G6PD inhibited. However, when
6NADPH ⫹ 6H⫹ ⫹ 3CO2 demand for NADPH increases, its utilization also
increases resulting in the fall of the intracellular NADPH
concentration. This removes the inhibition on the enzyme
The 1st phase of HMP shunt is the oxidative pathway in activity. Consequently, this reaction is speeded up, result-
which glucose 6-phosphate is oxidized and decarboxyl- ing in enhanced production of the required NADPH.
ated to produce two NADPH, a carbon dioxide and a
ribulose 5-phosphate. The interconversion phase consists
of reversible rearrangements of the phosphorylated D. Functions
pentoses. It links ribulose 5-phosphte to the glycolytic
pathway.
The pentose phosphate pathway is a multifunctional
pathway, unique in generation of two important prod-
ucts: pentoses and NADPH.
C. Regulation
Pentoses: The most important pentose is ribose
The regulatory enzymes of the pentose phosphate path- 5-phosphate, which is required for a number of bio-
way are glucose 6-phosphate dehydrogenase and 6-phospho- synthetic and other functions, as mentioned earlier in
gluconate dehydrogenase. Synthesis of both the enzymes is this chapter.
induced by insulin. Thus, in fed state, their intracellular NADPH: It has the same redox potential as NADH,
concentrations rise, leading to enhanced oxidation of but the functions of the two coenzymes are differ-
glucose through this pathway, and conversely in starva- ent. Whereas NAD⫹ collects hydrogen from catabolic
tion and diabetes mellitus, the pathway is suppressed. substrates for transfer to respiratory chain, NADPH is
Metabolism of Carbohydrates II: Secondary Pathways and Regulation of Blood Glucose Level 199
required for reductive biosynthesis and several other into the protective reduced form. This restores the level
functions: of reduced glutathione, which is oxidized while detoxify-
ing peroxides and other oxidizing agents. Thus, require-
Reductive biosynthesis: The synthesis of fatty acids and
ment of NADPH is relatively higher in erythrocytes. The extra
cholesterol from acetyl CoA require reductive power of
demand is met by a slight modification in the HMP-
NADPH.
shunt, as follows.
Antioxidant role: NADPH is necessary for the mainte-
nance of a reducing environment inside the cell. This is HMP-shunt can be Modified
necessary for avoiding damage to unsaturated fatty acids, Based on demand of the tissue, the HMP shunt is modi-
proteins and DNA by peroxides or other oxidizing agents, fied as below:
to which the cell is continuously exposed. NADPH plays
(a) Extra demand for NADPH: In erythrocytes require-
a key role in antioxidant defenses by converting the oxi-
ment for NADPH is relatively more, which is met as
dized glutathione into the reduced glutathione, which is
below:
protective:
Conversion of the pentoses to hexoses through the
H2O (GSSG)2 NADPH + H +
(oxidized) reversible Stage II reactions is favoured.
Glutathione Glutathione
The hexose phosphates so formed are then funneled
peroxidase reductase
into another cycle of oxidative reactions to produce
2GSH NADP +
H2O2 more of NADPH.
(reduced)
As diagrammed above, the reduced glutathione (GSH) This establishes a cyclic pathway, with each of these
detoxifies hydrogen peroxide; the enzyme glutathione per- cycles generating 2 NADPH molecules.
oxidase catalyzes this reaction. However, GSH is oxidized Stage II
during the reaction, and must be regenerated to continue Hexose phosphate Pentose phosphate
Stage I
detoxification. NADPH participates in the reaction
responsible for the regeneration of reduced glutathione
Generation of NADPH
from the oxidized one; glutathione reductase catalyzes this
reaction. (b) Extra demand for pentoses: Demand for pentoses is
greater than that for NADPH in some tissues. The non-

oxidative reactions can provide the ribose 5-phosphate
NADPH maintains the glutathione in reduced state by con-
verting the dimeric, oxidized glutathione (GSSG)2 into the molecules directly from fructose 6-phosphate via the
protective reduced form (GSH). This recycling of GSSG to Stage II reactions in such tissues.
GSH is crucial for maintaining antioxidant defense. Hexose phosphate Pentose phosphate

Phagocytosis (literally “cell eating”): It is the engulfment of Depending on cellular needs, ribulose 5-phosphate
solid particles into the cell, such as granulocytes, mono- is converted (via ribose 5-phosphate and xylulose 5-
cytes and macrophages. These cells participate in our phosphate) to hexose phosphates, which can re-enter into
immune system by eating up and destroying infectious another cycle of oxidative reactions to produce NADPH.
microorganisms (Chapter 33). NADPH is required for Conversely, hexose to pentose conversion is favoured
production of superoxide anion radicals by macro- in tissues having more demand for pentose.
phages, which have bactericidal activity (Case 27.1).
Metabolism of xenobiotics: Lipophilic xenobiotics are

metabolized to water-soluble products by hydroxylation Glucose 6-phosphate Dehydrogenase Deficiency
in the microsomal cytochrome P450 system; the process Some persons have a genetic defect in this enzyme, typi-
requires NADPH (Chapter 15). cally yielding an unstable enzyme, that has shorter half-
life in the RBC, or an enzyme that is unusually sensitive
Special function in erythrocytes: NADPH is especially to inhibition by NADPH. In either case, insufficient flux
important to prevent oxidative damage to the RBC mem- of the HMP-shunt and so decreased production of
brane and intracellular proteins in erythrocytes because NADPH results, and the cell’s ability to recycle GSSG to
the cell cannot replace these by new synthesis during its GSH is impaired. Drug-induced oxidative stress leads to
120-day lifespan. NADPH maintains the antioxidant lysis of RBCs (haemolysis); haemolytic anaemia is the
defenses by converting the oxidized, dimeric glutathione obvious consequence (Case 10.1).
UDP-Glucose
Glucose 6-phosphate dehydrogenase deficiency causes H2O + 2NAD+

drug induced haemolytic anaemia because of lack of UDP-Glucose dehydrogenase
NADPH, which maintains glutathione in its reduced state. 2H+ + 2NADH+
H2O UDP
Wernicke-Korsakoff Syndrome UDP-Glucuronate D-Glucuronate
Thiamine deficiency in alcoholics does not cause beri-

Glucuronate reductase
beri but Wernicke-Korsakoff syndrome. Reduced activity
of the thiamine-dependent transketolase enzyme occurs L-Gulonate
in this condition resulting is impairment of HMP shunt.
This condition is diagnosed by clinical features that Aldonolactonase
H 2O
include mental derangements, delirium and motor
incoordination, which may progress to chronic stage L-Gulonolactone
(called Korsakoff psychosis) characterized by amnestic syn-
Gulonolactone oxidase
drome. Early diagnosis and immediate treatment is 2H
essential, because brain damage in this condition is irre-
L-Ascorbic acid
versible.
Fig. 10.4. Reactions of uronic acid pathway.
This pathway is used by all plants and animals for the

II. Uronic Acid Pathway synthesis of vitamin C. However, some primates, such as
humans, monkeys, guinea pigs, as well as some birds and
Uronic acid pathway is a source of glucuronic acid. It also
fishes lost the capability to do so during evolution. This is
produces ascorbic acid in some animals. Glucuronic acid
because of genetic deficiency of the enzyme gulonolactone oxi-
is a useful product required in the synthesis of muco-
dase, which causes conversion of L-gulonolactone to L-ascorbic
polysaccharides, detoxification of some drugs and for
acid. It appears that the capacity to synthesize the ascorbic
conjugation of bilirubin and steroid hormones. The
acid was lost by these species because of a mutation, which
unutilized glucuronic acid (glucuronate) is converted to
was not lethal. These species require vitamin C in diet.
xylulose 5-phosphate, which is metabolized via the pen-
tose phosphate pathway.
A single enzyme defect in the uronic acid pathway
(deficiency of gulonolactone oxidase) causes inability of
A. Reactions of the Pathway primates to synthesize ascorbic acid.
Glucose is first converted to UDP-glucose by a series of

reactions, as discussed earlier in glycogenesis. The UDP B. UDP-Glucuronate is Metabolically
glucose further proceeds as below:
Active Compound
1. The glucose portion of the UDP-glucose is enzymatically
oxidized, resulting in formation of UDP-glucuronate. UDP-Glucuronate can readily denote the glucuronate
The reaction is catalyzed by the enzyme UDP-glucose residue for the following functions:
dehydrogenase. UDP-glucuronate is the metabolically 1. To detoxify foreign compounds or drugs. During detoxifica-
active compound, as discussed later. tion, the glucuronate residues are covalently attached to
2. From the UDP-glucuronate release of D-glucuronate these substances. Since glucuronate residues are strongly
occurs (Fig. 10.4). polar, their attachment imparts polar character to
3. D-Glucuronate is reduced by the NADPH-dependent these compounds, thus making possible their renal
enzyme glucuronate reductase to form L-gulonate. excretion. Bilirubin and steroid hormones are also
Glucuronate contains an acid carboxylate group at rendered more polar for excretion in this manner.
C-6, whereas gulonate contains this group at C-1.
4. L-Gulonate then loses a water molecule to form
Uronic acid pathway, a minor pathway of carbohydrate
L-gulonolactone. The enzyme is aldonolactonase.
metabolism, is a source of glucuronic acid for conjuga-
5. Removal of a pair of hydrogen atoms from the
tion of several endogenous and exogenous substances
L-gulonolactone by the enzyme gulonolactone oxidase
before excretion as glucuronides in urine and bile.
yields L-ascorbate or vitamin C.

Glycolysis, gluconeogenesis, glycogen metabolism, TCA cycle, and PPP

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Glycolysis, gluconeogenesis, glycogen metabolism, TCA cycle, and PPP

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METABOLISM OF CHAPTER

Glycolysis can be divided into two main stages. The stage

Reaction 7: ATP Production from 1,3-Bisphosphoglycerate C O P O− C OH C O

2. Reduction to lactate: Lactate is produced by glycolysis

B. Fate of Pyruvate Pyruvate Lactate

Glucose Acetyl CoA

charge stimulates it; Box 9.3), and by a number of posi-

cAMP Insulin Feedback

Aldolase B is an isoenzyme of the aldolase (see Reaction

catalyzes phosphorylation of fructose at C-1 to form ADP

“reducing sugar” (fructose) is present. Restriction of Hypertriglyceridaemia: Fructose is rapidly metabolized to

Glucose 6-phosphate Glycolysis

B. Feeder Pathways for Disaccharides

Metabolism of Amino Sugars

Lipids Carbohydrates Proteins

Fatty acids Glycerol Amino acids

Acetyl CoA mitochondrial membrane (Chapter 14). This arrange-

H2O COO− Oxaloacetate CH2

Fig. 9.16. Reactions of tricarboxylic acid (Krebs) cycle.

PFK1 Fatty acid

condenses with another molecule of acetyl CoA to initi- Glucose

The following points about TCA cycle are noteworthy: 4 CO2

E. Synthetic Functions of the TCA Cycle

Fig. 9.20. Regulation of pyruvate dehydrogenase complex (PDH).

Glycolysis modulators of the enzyme. Raised levels of these

Regulation of the Cyclic Pathway (Fig. 9.21)

gluconeogenesis. The pathway is important because

B. The Bypass Reactions of 1. Transport of pyruvate into mitochondrial matrix: Pyruvate

Inner mitochondrial membrane

yield glucose via the gluconeogenic sequence, as dis-

In addition to the three bypass steps, there are seven

F. Regulation inhibited by ATP, these two pathways are regulated in

Regulation of Pyruvate to Oxaloacetate Hormonal Regulation

ADP − Fructose 1,6-bis-P

1. As shown in Figure 9.26, compounds reﬂecting high PEP

CH2OH ATP ADP H2C O P

CH2OH CH2OH CH2OH

Reaction 3 Elongation of Chains by Glycogen Synthase

(a) (b) (c) (d)

α-1,6 bond α(1→6)-

Fig. 9.30. Glycogen degradation. Designation of glucose residues as in Figure 9.28.

Fate of Glucosyl Units Released from Glycogen Covalent Modulation

AMP Glucose 6- P ATP ADP Glucose

Glycogen phosphorylase b Glycogen phosphorylase a

Protein phosphatase-1 Phosphorylated enzyme

(Fig. 9.32). Thus, it is activated by protein kinase and ren-

Regulation by Calcium Ions

Protein kinase Glucose 6- P

ATP cAMP + PPi

Protein kinase (inactive) Protein kinase (active)

+ − Protein phosphatase. The latter causes dephosphorylation of gly-

is that a single hormone molecule can generate thousands of

D. Glycogen Storage Diseases Table 9.6. Glycogen storage diseases

von Gierke’s disease (Type I): It is a classical hepatic gly-

Her’s disease (Type VI): Liver glycogen stores cannot be Exercises

CASE 9.2 An unconscious 5-year old child

Fig. 10.1. Overview of the pentose phosphate pathway.

CH2OH NADPH + H+ NADP+

Fig. 10.2. The oxidative reactions of pentose phosphate pathway.

Xylulose-5-P Ribose-5-P Xylulose-5-P