a brief introduction and metabolic fate of glycolysis

1. GLYCOLYSIS
Presented by:
Dipesh Tamrakar
Msc. Clinical Biochemistry

2. Overview
• Introduction
• History
• Steps of glycolysis
• Regulation of glycolysis
• Biomedical importance of glycolysis
• Glycolysis in Cancer cells
• Fates of pyruvate
• Feeder pathways for glycolysis

3. Definition
• Glykys = Sweet, Lysis = splitting (Greek)
• A molecule of glucose is degraded in a series of enzyme-
catalyzed rxn to yield 2 molecules of 3C pyruvate
• During the sequential rxn of glycolysis, some of the free energy
released from glucose is conserved in the form of ATP and NADH
• Glycolysis was the first metabolic pathway to be elucidated
• The glycolytic breakdown of glucose is the sole source of
metabolic energy in some mammalian tissues and cell types
(erythrocytes, renal medulla, brain, and sperm)

4. • Glycolysis occurs in most organisms in the cytosol of the cell.
• The most common type of glycolysis is the Embden–Meyerhof–Parnas
(EMP pathway), which was discovered by Gustav Embden, Otto
Meyerhof, and Jakub Karol Parnas
• Glucose first phosphorylated irreversibly by hexokinase and traps the
sugar as cytosolic Glucose-6-P & commits it to further metabolism in
the cell
4

5. • Phosphate esters are charged, hydrophilic compounds that do not
readily penetrate cell membranes & glycolytic enzymes are present in
cytosol
• Hexokinase IV also serves as a glucose sensory in hypothalamic
neurons, playing a key role in the adrenergic response to
hypoglycemia
• In beta cells, glucokinase functions as a glucose sensors, determining
the threshold for insulin secretion
5

6. History of glucose metabolism
Year in AD Scientists Discovery
1768 Abbate Spallanzani Showed that living things take up
O2 and give up CO2
1860 Louis Pastueur Demonstrated fermentation of
glucose to alcohol by yeast
1893 Gad Found out that lactic acid is
formed during muscle
contraction
1902 Sir Walter Fletcher Lactic acid is derived from
glycogen

7. 1914 Gustav George
Embden
Studied LA formation from
pyruvate
1915 Von Euler Chelpin First identified Hexokinase
activity
1919 Otto Fritz Meyerhof Enunciated most of the steps of
glycolytic pathways
1920 Sir Arthur Harden Phosphofructokinase
1934 Parnas Pyruvate kinase
1935 Mayerhof enolase
1920-
1943
Many discovers All enzymes of glycolytic
pathway has been identified
and characterized

8. Fates of glucose

9. Overview of glycolysis
1. Preparatory phase (the energy requiring stage):
• One molecule of glucose is converted into two molecules of
glyceraldhyde-3-phosphate
• Requires 2 molecules of ATP (energy loss)
• Glucose is first phosphorylated at the hydroxyl group on C-6
• The D-glucose 6 phosphate thus formed is converted to D-
fructose 6 phosphate
• Phosphorylated at C-1 yield D-fructose 1,6-bisphosphate
• Spliting fructose 1,6-bisphosphate yield two 3-C molecules:
dihydroxyacetone phosphate and glyceraldehyde 3-
phosphate

10. 2. Payoff phase (the energy producing stage(:
• 2 molecules of glyceraldehyde-3-phosphate are converted
into pyruvate (aerobic glycolysis) or lactate (anaerobic
glycolysis)
• This stage produce ATP molecules (energy production)
• Glyceraldehyde 3-phosphate is oxidized and phosphorylated
by inorganic phosphate to from 1,3-bisphoglycerate
• Energy is then released as the 2 molecules of 1,3-
bisphosphoglycerate are converted to 2 mol of pyruvate
• Energy is also conserved in the pay-off phase in the formation
of 2 mol of the electron carrier NADH per mol of glucose

14. • Irreversible, catalyzed by hexokinase
• Hexokinase also catalyzes the phosphorylation of other common
hexoses s/a fructose and mannose
• Requires Mg2+ for its activity (true substrate is MgATP2- complex

15. Function of Mg2+
• Mg shields the negative charges of the phosphoryl groups in
ATP, making the terminal phosphorus atom an easier target for
nucleophilic attack by an –OH of glucose
• Reaction is accompanied by considerable loss of free energy as
heat, and hence under physiologic conditions is regarded as
irreversible.

16. Hexokinase
• Hexokinase undergoes an induced fit by change in shape when
it binds glucose
• 4 isoenzymes (I,II,III & IV) encoded by 4 different genes
• Predominant hexokinase isoenzyme of myocytes (hexokinase II)
has high affinity for glucose (1/2 saturation at 0.1 mM)
• Muscle hexokinase I and II are allosterically inhibited by their
product, glucose 6 phosphate, so whenever the cellular
concentration of glucose 6 phosphate rises above its normal
level, these are temporarily and reversibly inhibited

17. Regulation of glycolysis by hexokinase
• The different hexokinase isozymes of liver and muscle reflect
the different roles of these organs in glucose metabolism
• Immediately after a carbohydrate-rich meal, when blood
glucose is high, glucose enters hepatocytes via GLUT2 and
activates hexokinase IV
• During fast, fructose-6-phosphate triggers the inhibition of
hexokinase IV by regulatory protein. So liver does not
compete with other organs for the scarce glucose
• Hexokinase IV is not inhibited by glucose-6-phosphate and
can therefore continue to operate when the accumulation of
glucose-6-phosphate completely inhibits hexokinases I-III

18. Regulation of glucokinase activity by glucokinase regulatory
protein
• Inhibited by fructose 6-phosphate
• Regulation achieved by reversible
binding to the hepatic protein
glucokinase regulatory protein (GKRP)
• When blood glucose level increases,
glucokinase is released from GKRP
and enzyme reenters the cytosol and
glycolysis is proceed

19. Hexokinase Glucokinase
Occurrence In all tissues Only in liver
Km value 10-2 mmol/L 20 mmol/L
Affinity to substrate High Low
Specificity Acts on glucose, fructose
and mannose
Acts only on glucose
Induction Not induced Induced by insulin and
glucose
Function Even when blood sugar
level is low, glucose is
utilized by body cells
Acts only when blood
glucose level is more
than 100 mg/dl; then
glucose is taken up by
liver cells for glycogen
synthesis

20. • Phosphohexose isomerase catalyzes the reversible rxn
• This rxn involves an enediol intermediate

21. The phosphohexose isomerase reaction

22. • Phosphofructokinase-1 (PFK-1) catalyzes the transfer of a
phosphoryl group from ATP to fructose 6-phosphate to yield
fructose 1,6-bisphosphate
• PFK 1 activity is increased whenever the cells ATP supply is
depleted or when the ATP breakdown products, ADP and AMP
accumulated

23. Regulation of PFK-1
• PFK-1 is different from PFK-2 that catalyzes formation of
Fructose 2,6-BP
• PFK is the rate limiting enzyme & control point of glycolysis
PFK-l is subject to complex allosteric regulation:
Regulation by energy levels within the cell:
• PFK-1 activity increased   AMP ,  ADP ,  citrate
• PFK-1 activity inhibited   ATP,  Citrate

24. Regulation of PFK-1 Contd..
Regulation by fructose 2,6-bisphosphate:
• the most potent activator and works even at high ATP
• also acts as an inhibitor of Fructose 1,6-bisphosphatase
(gluconeogenesis)
• Fructose 2,6-bisphosphate is formed by phosphorylation of
Fructose 6-P by enzyme PFK-2 and is broken down by FBPase-2
• The cellular concentration of fructose 2,6-bisphosphate is
regulated by insulin and glucagon

25. Regulation of fructose 2,6-bisphosphate level

26. • Effect of elevated insulin concentration on the intracellular
concentration of fructose 2,6-bisphosphate in liver

27. • Glucagon, via cAMP pathway, activates cAMP dependent
protein kinase. Phosphorylation of this protein enhances its
FBPase-2 activity and inhibits its PFK-2 activity. It, therefore,
lowers the concentration of fructose 2,6-BP inhibiting
glycolysis and stimulating gluconeogenesis
• Insulin has the opposite effect, stimulating the activity of a
phosphoprotein phosphatase that catalyzes removal of
phosphate group from the bifunctional protein, enhancing its
PFK-2 activity
• Xylulose 5-P, a product of PP pathway, is another regulator
Regulation of PFK-1 Contd..

28. • Fructose 1,6 bisphosphate aldolase ( aldolase) catalyzes a
reversible aldol condensations
• Fructose 1,6 bisphosphate is cleaved to yield 2 different
triose phosphates, glyceraldehyde 3-phosphate (aldose) and
dihydroxyacetone phosphate (ketose)

29. • Only glyceraldehyde 3-phosphate can be directly degraded in
the subsequent steps of glycolysis
• Triose phosphate isomerase converts dihydroxy acetone
phosphate

30. • The first step in the payoff phase
• Oxidation of glyceraldehyde 3-phosphate to 1,3-
bisphoglycerate catalyzed by Glyceraldehyde 3-Phosphate
Dehydrogenase

32. Rapaport-Leubering cycle:
• In RBC some of the 1,3-BPG is converted to 2,3-BPG by the
action of bisphosphoglycerate mutase
• Significance of 2,3-BP:
1. Allows glycolysis to proceed without the synthesis of ATP. So
very useful when RBC has no requirement of energy.
2. 2,3-BPG binds to Hb, decreasing its affinity for oxygen and so
making oxygen more readily available to tissues.
3. 2,3-BPG synthesis increases in hypoxic condition, high
altitude, fetal tissue, anemic condition etc..

33. • Phosphogylcerate kinase transfers
the high-energy phosphoryl group
from the carboxyl group of 1,3-
bisphosphoglycerate to ADP
forming ATP and 3-
phosphoglycerate
• The formation of ATP by phosphoryl
group transfer from a substrate
such as 1,3-bisphosphoglycerate is
referred to as a substrate-level
phosphorylation

34. • The enzyme phosphoglycerate mutase catalyzes a reversible
shift of the phosphoryl group between C2 and C3 of glycerate
• Mg2+ is essential for this reaction
• Reaction occurs in 2 steps

35. The phosphoglycerate
mutase reaction

36. • Dehydration of 2-phosphoglycerate to phosphenolpyruvate
• Generated a compound with high phosphoryl group transfer
potential
• Enolase promotes reversible removal of water molecule from 2-
phosphoglycerate

37. • Last step in glycolysis is the
transfer of the phosphoryl group
from phosphoenolpyruvate to
ADP
• Catalyzed by pyruvate kinase
which requires K+ and either
Mg2+ or Mn2+
• Pyruvate first appears in enol
form then tautomerize to its keto
form

38. Pyruvate kinase is allosterically inhibited by ATP
• High concn of ATP, acetyl-CoA and long chain fatty acids
allosterically inhibits all isoenzymes of pyruvate kinase
• The liver isozymes (L-form), but not the muscle isozymes is
subject to further regulation by phosphorylation

39. Overall energy balance sheet
Glucose + 2ATP + 2NAD + 4ADP + 2Pi
2Pyruvate + 2ADP +2NADH +2H+ + 4ATP + 2H2O
This accounts for :
1. Fate of carbon skeleton of glucose
2. The input of Pi and ADP and output of ATP
3. The pathway of electrons in the oxidation-reduction
reactions
Glucose + 2NAD + 2ADP + 2Pi
2Pyruvate +2NADH +2H+ + 2ATP + 2H2O
(under aerobic condition)

41. Factors Regulating Glycolysis
1. Hexokinase :
• high affinity for glucose will phosphorylate glucose even at
low glucose conc to provide energy to tissues that depend on
glycolysis for energy needs (brain, RBCs)
• G-6-phosphate has feedback inhibitory effect on the enzyme
2. Glucokinase :
• Low affinity and high km for glucose is present only in tissues
where the phosphorylation has to take place when glucose is
available in plenty
• In the liver glucokinase phosphorylates glucose which can be
used for glycogen synthesis.

42. • In the beta cells of pancreases, the glucokinase
phosphorylates glucose when the intracellular conc is
sufficiently high so that ATP is produced within the cells
• Insulin also induces glucokinase
3. Phosphofructokinase (PFK) is the most important rate-
limiting enzyme for glycolysis pathway
4. ATP and citrate are the most important allosteric inhibitors
5. AMP acts as an allosteric activator
6. Fructose-2,6-bisphosphate increases the activity of
phosphofructokinase

43. 7. Pyruvate kinase catalyzes an irreversible step and is a
regulatory enzyme of glycolysis. When energy is in plenty
in the cell, glycolysis is inhibited. Insulin increases its
activity whereas glucagon inhibits. Pyruvate kinase is
inactive in the phosphorylated state.
8. Insulin favors glycolysis by activating the key glycolytic
enzymes
9. Glucocorticoids inhibit glycolysis and favors
gluconeogenesis
PASTEUR EFFECT
Under aerobic condition, glycolysis is inhibited by oxygen
which is known as Pasteur effect

44. Importance of phosphorylated intermediate
1. Because the plasma membrane generally lacks transporters
for phosphorylated sugars, the phosphorylated glycolytic
intermediates cannot leave the cell.
2. Phosphoryl groups are essential components in the enzymatic
conservation of metabolic energy
3. Binding energy resulting from the binding of phosphate
groups to the active sites of enzymes lowers the activation
energy and increases the specificity of the enzymatic
reactions

45. Inhibition of Glycolysis
• Inhibitors of Glycolysis; Sulfhydryl reagents and fluoride
• Glyceraldehyde 3-Phosphate dehydrogenase is vulnerable to
inhibition by sulfhydryl reagents s/a iodoacetate
• Fluoride is a potent inhibitor of enolase
• Mg2+ and Pi form an ionic complex with fluoride ion; responsible by
interfering with binding of enolase substrate

46. Importance of glycolysis by birth:
• Except brain, circulation of blood decreases to most parts of the body
of the neonate during delivery
• The brain is not normally deprived of O2 during delivery, but other
tissues must depend on glycolysis for their supply of ATP until
circulation returns to normal & O2 becomes available again
• This converses O2 for use by the brain & assure survival of brain tissue
in the time of stress
47

47. • RBC lacks mitochondria & therefore are unable to convert pyruvate to
CO2 and H2O
• The cornea, lens & regions of the retina have a limited blood supply &
also lack mitochondria and depend on glycolysis as the major
mechanism for ATP production
• Kidney medulla, testis, leukocytes & white muscle fibers are almost
totally dependent on glycolysis as a source of ATP because they have
relatively very few mitochondria
• Brain consumes about 120 gm of glucose every day whereas other
tissues dependent primarily on glycolysis for ATP production consume
about 40 gm of glucose per day in a normal adult
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48. Deficiency of Glycolytic enzymes
Hexokinase : Haemolytic anaemia
Phosphofructokinase: Glycogen Storage Disease Type
VII or Tarui's disease; muscular metabolic disorder
pyruvate kinase : Haemolytic anaemia
• Inactivating mutations of glucokinase are the cause of rare form
of diabetes, maturity onset diabetes of the young type 2 (MODY
2)
• It is characterized by impaired insulin secretion and
hyperglycemia
49

49. • Glucose uptake and glycolysis proceed about 10 times faster in most
solid tumors than in non-cancerous tissues
• Tumor cells commonly experience hypoxia so depend on anaerobic
glycolysis
• To compensate high ATP need from glycolysis, some tumor cells
overproduce several glycolytic enzymes
• Hypoxia-inducible transcription factor (HIF-1) is protein that acts at the
mRNA level to synthesis atleast 8 glycolytic enzymes
• This gives tumor cells the capacity to survive in hypoxia until the supply
of blood vessels has caught up tumor growth
Glucose catabolism in cancerous tissues
50

50. • The increase in glycolytic activity ultimately counteracts the effects of
hypoxia by generating sufficient ATP from this anaerobic pathway
• This phenomenon was first described in 1930 by Otto Warburg and is
referred to as the Warburg effect.
• The Warburg hypothesis claims that cancer is primarily caused by
dysfunctionality in mitochondrial metabolism, rather than because of
uncontrolled growth of cells.
• A number of theories have been advanced to explain the Warburg
effect.
• One such theory suggests that the increased glycolysis is a normal
protective process of the body and that malignant change could be
primarily caused by energy metabolism.
51

51. • The high glycolysis rate has important medical applications, as high
aerobic glycolysis by malignant tumors is utilized clinically to diagnose
and monitor treatment responses of cancers by imaging uptake of
2-18F-2-deoxyglucose (FDG)
(a radioactive modified hexokinase substrate) with positron emission
tomography(PET).
• There is ongoing research to affect mitochondrial metabolism and
treat cancer by reducing glycolysis and thus starving cancerous cells in
various new ways, including a ketogenic diet.
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52. 53

53. 54

54. • The Histone Deacetylase SIRT6 Is a
Tumor Suppressor that Controls
Cancer Metabolism
• SIRT6 can lead to tumor formation
even in the absence of oncogene
activation
• Inhibition of glycolysis in SIRT6-
deficient cells abrogates tumor
formation
55

55. Pyruvate kinase
• Pyruvate kinase enzyme catalyzes the last step of glycolysis
• Pyruvate kinase catalyzes the transfer of a phosphate
group from phosphoenolpyruvate (PEP) to ADP, yielding one molecule
of pyruvate and one molecule of ATP.
• Liver pyruvate kinase is indirectly regulated by epinephrine and glucagon,
through protein kinase A.
• This protein kinase phosphorylates liver pyruvate kinase to deactivate it.
56

56. • Muscle pyruvate kinase is not inhibited by epinephrine activation of
protein kinase A.
• Glucagon signals fasting (no glucose available). Thus, glycolysis is inhibited
in the liver but unaffected in muscle when fasting.
• An increase in blood sugar leads to secretion of insulin, which activates
phosphoprotein phosphatase I, leading to dephosphorylation and
activation of pyruvate kinase.
• These controls prevent pyruvate kinase from being active at the same
time as the enzymes that catalyze the reverse reaction (pyruvate
carboxylase and phosphoenolpyruvate carboxykinase), preventing a futile
cycle.
57

57. Pyruvate Kinase Deficiency
• Pyruvate kinase deficiency is an inherited metabolic disorder of the
enzyme pyruvate kinase which affects the survival of red blood cells
• With insufficient ATP in an erythrocyte, all active processes in the cell come
to a halt.
• Sodium potassium ATPase pumps are the first to stop. Since the cell
membrane is more permeable to potassium than sodium, potassium leaks
out.
• Intracellular fluid becomes hypotonic, water moves down its concentration
gradient out of the cell. The cell shrinks and cellular death occurs, this is
called 'dehydration at cellular level’.
• This is how a deficiency in pyruvate kinase results in hemolytic anaemia
58

58. Cause
• Pyruvate kinase deficiency is due to a mutation in the PKLR gene.
• There are four pyruvate kinase isoenzymes, two of which are encoded
by the PKLR gene (isoenzymes L and R, which are used in the liver
and erythrocytes, respectively).
• 180 different mutations have been found on the gene coding for the L
and R isoenzymes, 124 of which are single-nucleotide missense
mutations.
59

59. Regulation of pyruvate kinase 60

60. Fates of pyruvate 61

61. • Pyruvate is oxidized, with loss of its carboxyl group as CO2 to yield the
acetyl group of acetyl-coenzyme A
• The acetyl group is then oxidized completely to CO2 by the citric acid
cycle
• The electrons from these oxidations are passed to 02 through a chain
of carriers in mitochondria, to form H2O.
• The energy from the electron-transfer reactions drives the synthesis
of ATP mitochondria
Fate of pyruvate under aerobic condition: Acetyl Co A
62

62. • The second route for pyruvate is its reduction to lactate via lactic acid
fermentation.
• When vigorously contracting skeletal muscle must function under low
oxygen conditions (hypoxia), NADH cannot be reoxidized to NAD+,
but NAD+ is required as an electron acceptor for the further oxidation
of pyruvate.
• Under these conditions pyruvate is reduced to lactate, accepting
electrons from NADH and thereby regenerating the NAD+ necessary
for glycolysis to continue.
• Certaint issues and cell types (retina and erythrocytes, for example)
convert glucose to lactate even under aerobic conditions, and lactate
is also the product of glycolysis under anaerobic conditions in some
microorganisms
Fate of pyruvate under anaerobic condition: Lactate
63

63. Lactic acidosis
• Elevated concentrations of lactate in the plasma occur d/t increased
production or decreased utilization.
• Mild lactic acidosis: strenuous exercise, shock, respirstory diseases,
cancer, low pyruvate dehydrogenase activity and Von Gierke’s disease.
• Severe lactic acidosis : MI, pulmonary embolism, uncontrolled
haemorrhage and severe shock
• The failure to bring adequate amounts of oxygen to the tissues results
in impaired oxidative phosphorylation and decreased ATP synthesis.
• To survive, the cells use anaerobic glycolysis as a backup system for
generating ATP, producing lactic acid as the end-product
64

64. • The third major route of pyruvate catabolism leads to ethanol.
• In some plant tissues and in certain invertebrates, protists and
microorganisms such as brewer’s or baker's yeast, pyruvate is converted
under hypoxic or anaerobic conditions to ethanol and CO2, a process
called ethanol (alcohol) fermentation
• Fermentation, which extract energy (as ATP) but do not consume O2 or
change the concentration of NAD+ or NADH.
• It can, for example provide the carbon skeleton for the synthesis of the
amino acid alanine or for the synthesis of fatty acids
Fate of pyruvate under anaerobic condition: Fermentation
65

65. Fermentation
• 1st step: Pyruvate is decarboxylated to acetaldehyde by pyruvate
decarboxylase, requires Mg2+ and has a tightly bound co-enzyme
TPP(Thiamine pyrophosphate)
• 2nd step: Acetaldehyde is reduced to ethanol by alcohol
dehydrogenase utilizing NADH previously produced during glycolysis
66

66. •The overall reaction is:
•Pyruvate decarboxylase is present in brewer’s and baker’s yeast
(Saccharomyces cerevisiae), CO2 produced by PD responsible for
carbonation .
•The enzyme is absent in vertebrates and other organisms that carry out
lactic acid fermentation
•TPP, the coenzyme form of vitamin B1 , carries 2 of the 3 carbons of
pyruvate in the form of a hydroxyethyl group, which is subsequently
released as acetaldehyde
67

67. • Alcohol dehydrogenase is present in
many organisms that metabolize
ethanol, including humans.
• In the liver it catalyzes the oxidation of
ethanol, either ingested or produced by
intestinal microorganisms with the
concomitant reduction of NAD+ to
NADH.
• In this case, the reaction proceeds in the
direction opposite to that involved in the
production of ethanol by fermentation
68

68. Application of Fermentation
• Used in production and preservation of foods
• Fermentations are used to produce some common foods eg. Yogurt
(Lactobacillus bulgaricus), Swiss cheese (Propionibacterium
freudenreichii), kimchi, pickles, sausage, soy sauce etc.
• The drop in pH associated with fermentation also helps to preserve the
food
• Fermentation process is also used to produce various industrial
chemicals s/a formic acid, succinic acid, glycerol, methanol, butanol,
ethanol, etc.
69

69. • Many carbohydrates besides glucose meet their catabolic fate in
glycolysis after being transformed into one of the glycolytic
intermediates:
1. Glycogen and starch.
2. The Disaccharides maltose, lactose, trehalose, and sucrose.
3. The monosaccharides fructose, mannose, and galactose.
Feeder Pathways for Glycolysis
70

70. 71

71. Glycogen and starch:
• glycogen phosphorylase (starch phosphorylase in plants)
enzymes catalyze an attack by Pi on the (α14) glycosidic linkage that
joins the last two glucose residues at a nonreducing end, generating
glucose 1-phosphate and a polymer one glucose unit shorter.
• Starch glycogen phosphorylase acts repetitively until it approaches an
(α16) branch point
• debranching enzyme removes the branches.
Phosphoglucomutase:
72

72. Dietary Polysaccharides and Disaccharides Undergo Hydrolysis to
Monosaccharides
73

73. FRUCTOSE: ENTRY INTO GLYCOLYTIC PATHWAY
Major pathway of fructose entry into glycolysis in the muscles and kidney
is catalysed by hexokinase
In the liver, however, Fructose enters by a different pathway. The liver
enzyme fructokinase catalyzes the phosphorylation of fructose at C-1
rather than C-6
74

74. • Fructose 1- phosphate is then cleaved to Glyceraldehyde and DHAP by
Fructose 1-phosphate aldolase
• DHAP is converted to Glyceraldehyde 3-phosphate by glycolytic enzyme
triose phosphate isomerase.
• Glyceraldehyde is phosphorylated by ATP and triose kinase to
glyceraldehyde 3-phosphate.
• Both products of fructose 1- phosphate enter glycolytic pathway as
glyceraldehyde 3-phosphate.
75

75. GALACTOSE UTILIZATION
Galactose passes in the blood from the intestine to the liver, where it is
first phosphorylated at C-1 at expense of ATP by enzyme galactokinase
• A defect in any of the three enzymes in this pathway causes
galactosemia in humans.
• In Galactokinase-deflciency Galactosemia, high galactose
concentrations are found in blood and urine. Affected individuals
develop cataracts in infancy, caused by deposition of the galactose
metabolite galactitol in the lens
• In Transferase-deficiency Galactosemia, poor growth in children, speech
abnormality, mental deficiency and liver damage can be observed
76

76. Conversion of galactose to glucose1 –phosphate
• The galactose l-phosphate is then converted to
its epimer at C-4, glucose 1-phosphate by UDP-
glucose:galactose 1-phosphate
uridylytransferase
• UDP-galactose is converted to UDP-glucose by
UDP-glucose 4-epimerase
• The epimerization involves first the oxidation of
the C-4 -OH group to a ketone by NAD+, then
reduction of the ketone to an –OH by NADH,
with inversion of the configuration at C-4.
77

77. ENTRY OF MANNOSE
• D-Mannose can be phosphorylated at C-6 by hexokinase
• Mannose 6-phosphate is isomerized by phosphomannose isomerase
to yield fructose 6-phosphate, an intermediate of glycolysis.
78

78. Mitochondrial poisons
1. Atovaquone is an antimalarial drug used for the treatment of
pneumocystis pneumonia and toxoplasmosis and is FDA approved.
At the molecular level it is a potent and selective inhibitor of
OXPHOS by targeting mitochondrial complex III, inducing aerobic
glycolysis and oxidative stress in cancer stem cells
2. Metformin; main mechanism of action is inhibition of
mitochondrial complex I, increasing the glycolytic pathway through
reduction of OXPHOS
3. Phenformin is metformin’s predecessor with similar effects on lactic
acid production, but is a more powerful inhibitor of the
mitochondrial respiratory chain which entails an increased risk of
lactic acidosis.
79

79. • Doxycycline is an antibiotic that exerts inhibition of mitochondrial
protein synthesis and reduces mitochondrial complex I activity
• All four pharmaceuticals described as mitochondrial poisons have in
common their inhibitory activity on OXPHOS and increase in lactic
acid production through increased aerobic glycolysis.
• Metformin, phenformin and doxycycline are weak inhibitors of
mitochondrial complex I and with no effect on the rest of the
mitochondrial complexes
80

80. PASTEUR EFFECT
• The inhibition of glycolysis by O2 is k/a Pasteur effect.
• Discovered by Louis Pasteur
• Pasteur effect is due to the inhibition of the enzyme
phosphofructokinase
• The levels of glycolytic intermediates from F1,6-bisphosphate
onwards decrease while the earlier intermediates accumulate
• More ATP is produced under aerobic conditions than under anaerobic
conditions, therefore less glucose is consumed aerobically
81

81. Crabtree effect
• Inhibition of oxygen consumption by the addition of glucose to tissues
having high aerobic glycolysis
• Basically, this is opposite to that of Pasteur effect
• Is due to increased competition of glycolysis for inorganic phosphate
and NAD+ which limits their phosphorylation and oxidation
82

82. Futile cycle
• A futile cycle, also known as a substrate cycle, occurs when
two metabolic pathways run simultaneously in opposite directions and
have no overall effect other than to dissipate energy in the form of heat
• For example, if glycolysis and gluconeogenesis were to be active at the
same time, glucose would be converted to pyruvate by glycolysis and then
converted back to glucose by gluconeogenesis, with an overall
consumption of ATP
• Futile cycles may have a role in metabolic regulation, where a futile cycle
would be a system oscillating between two states and very sensitive to
small changes in the activity of any of the enzymes involved.
83

83. • The cycle does generate heat, and may be used to maintain
thermal homeostasis, for example in the brown adipose tissue of
young mammals, or to generate heat rapidly, for example in insect flight
muscles and in hibernating animals during periodical arousal from torpor.
• It has been reported that the glucose metabolism substrate cycle is not a
futile cycle but a regulatory process. For example, when energy is suddenly
needed, ATP is replaced by AMP, a much more reactive adenine.
• The reciprocal actions of fructose 2,6-bisphosphate on glycolysis
(activation) and gluconeogenesis (inhibition) ensure that both pathways are
not fully active at the same time, preventing a futile cycle in which glucose
would be converted to pyruvate followed by resynthesis of glucose from
pyruvate
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84. Thank-you
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