Glycolysis
Gandham. Rajeev
Metabolism
Metabolism is “the entire set of enzyme-
catalyzed transformations of organic
molecules in living cells.
Two broad classes:
Catabolism & Anabolism
Catabolic Pathways:
Transform fuels into cellular energy
Anabolic Pathways
Requires inputs of energy to proceed.
Useful energy + small molecules
complex molecules.
Pathways that can be either anabolic or
catabolic, depending on the energy
conditions in the cell are referred to as
amphibolic pathways
Glycolysis
Glycolysis occurs in almost every living cell.
It was the first metabolic sequence to be
studied.
This pathway is also called Embden-
Meyerhof pathway (E.M-Pathway).
It occurs in cytosol.
Definition
Glycolysis is defined as the sequence of
reactions converting glucose to pyruvate or
lactate, with the production of ATP.
Salient features:
Takes place in all cells of the body.
The enzymes of this pathway are present in
the cytosomal fraction of the cell.
Glycolysis occurs in the absence of oxygen
(anaerobic) or in presence of oxygen
(aerobic).
Lactate is the end product under anaerobic
condition.
In aerobic condition, pyruvate is formed,
which is then oxidized to CO2 & H2O.
Glycolysis is a major pathway for ATP
synthesis in tissues lacking mitochondria,
erythrocytes, cornea, lens etc.
Glycolysis is very essential for brain which is
dependent on glucose for energy.
The glucose in brain has to undergo
glycolysis before it is oxidized to CO2 & H2O.
Glycolysis is a central metabolic pathway
with many of its intermediates providing
branch point to other pathways.
The intermediates of glycolysis are useful for
the synthesis of amino acids and fat.
Glucose entry into cells
Glucose transporter-4 (GluT4) transports
glucose from extracellular fluid to muscle
cells & adipocytes.
This is under the influence of Insulin.
In diabetes mellitus, insulin deficiency hinders
the entry of glucose into the peripheral cells.
GluT2 is the transporter in liver cell.
It is not under the control of insulin.
Reactions of Glycolysis
Divided into three distinct phases.
Energy investment phase or priming phase
Splitting phase
Energy generation phase.
Energy investment phase
Glucose obtained from
The diet through intestinal hydrolysis of
lactose, sucrose, glycogen, or starch is
brought into the hexose phosphate pool
through the action of hexokinase.
Free glucose is phosphorylated to glucose 6
phosphate by hexokinase
Hexokinase splits the ATP into ADP & Pi,
the Pi is added to the glucose.
Hexokinase Hexokinase is a key
glycolytic enzyme.
Hexokinase or Glucokinase
Glucose Glucose 6-Phosphate
Mg+2
ATP ADP
• Phosphorylated sugar molecules do not
readily penetrate cell membranes without
specific carriers, this commits glucose to
further metabolism in the cell.
• In all tissues, the phosphorylation of glucose
is catalyzed by hexokinase, one of the three
regulatory enzymes of glycolysis.
Hoxokinase and glucokinase
Hexokinase Glucokinase
Occurrence In all tissues Only in liver
Km Value 10-2 mmol/L 20 mmol/L
Affinity to substrate High Low
Acts on glucose, fructose
Specificity Acts only on glucose
and mannose
Induced by insulin &
Induction Not induced
glucose
Acts only when blood
Even when blood sugar glucose level is more then
Function level is low, glucose is 100 mg/dl; then glucose is
utilized by body cells taken up by the liver cells
for glycogen synthesis
Isomerization of Glucose 6-P
Glucose 6 P is a central molecule with a
variety of metabolic fates- glycolysis,
glycogenesis, gluconeogenesis and HMP
pathway.
The isomerization of Glucose 6-P (an aldose
sugar) to Fructose 6-P (a ketose sugar) is
catalyzed by phosphohexose isomerase
It requires Mg+2 ions.
The reaction is readily reversible, is NOT
a rate limiting or regulated step.
Phosphohexose isomerase & Mg+2
Glucose 6-Phosphate Fructose 6-Phosphate
Phosphorylation of Fructose 6-P
• Fructose 6- phosphate is phosphorylated to Fructose
1, 6- bisphosphate by Phosphofructokinase (PFK)
• The PFK reaction is the rate-limiting step.
• It is controlled by the concentrations of the
substrates ATP & Fructose 6-P
Phosphofructokinase
Fructose 6P Fructose 1, 6-bisPhosphate
Mg+2
ATP ADP
Splitting Phase
• The six carbon Fructose 1, 6- bisphosphate is split
to 2 three carbon compounds.
• Glyceraldehyde 3- phosphate & Dihydroxy acetone
phosphate by the enzyme aldolase (Fructose 1, 6-
bisphosphate aldolase).
• The reaction is reversible is not subject to regulation.
Fructose 1,6- Aldolase
Glyceraldehyde 3-
bisphosphate Phosphate + DHAP
Isomerization of DHAP
• Phosphotriose isomerase catalyzes the reversible
interconversion of dihydroxyacetone phosphate &
glyceraldehyde 3-phosphate.
• Two molecules of glyceraldehyde 3-phosphate are
obtained from one molecule of glucose.
Phosphohexose isomerase
DHAP Glyceraldehyde 3-Phosphate
Oxidation of glyceraldehyde 3P
Glyceraldehyde 3-phosphate dehydrogenase
converts Glyceraldehyde 3-phosphate to 1,3-
bisphosphoglycerate.
This step is important as it is involved in the
formation of NADH +H+ & a high energy
compound 1,3- bisphosphoglycerate.
In aerobic condition, NADH passes through
the ET C and 6 ATP are synthesized by
oxidative phosphorylation.
Glyceraldehyde 3P-
dehydrogenase
Glyceraldehyde 3P 1,3-bisphosphoglycerate
NAD NADH+H+
Pi
Formation of ATP from 1,3-
bisphosphoglycerate & ADP
• The enzyme phosphoglycerate kinase acts on
1,3- bisphosphoglycerate resulting in the
synthesis of ATP and formation of 3-
phosphoglycerate.
Phosphoglycerate kinase
1,3-bisphosphoglycerate 3P-glycerate
Mg +2
ADP ATP
This step is a substrate-level phosphorylation
Production of a high-energy P is coupled to
the conversion of substrate to product, instead
of resulting from oxidative phosphorylation.
The energy will be used to make ATP in the
next reaction of glycolysis.
• The formation of ATP by P group transfer
from a substrate such as 1,3-
bisphosphoglycerate is referred to as a
substrate-level phosphorylation.
• Unlike most other kinases, this reaction is
reversible.
3- Phosphoglycerate is converted to 2-
Phosphoglycerate by phosphoglycerate
mutase
This is isomerization reaction.
Phosphoglycerate mutase
3-Phosphoglycerate 2P-glycerate
The high energy compound PEP is generated
from 2- Phosphoglycerate by the enzyme
enolase.
This enzyme requires Mg+2 or Mn+2 and is
inhibited by fluoride.
Enolase
2-Phoglycerate Phosphoenolpyruvate
Mg+2
The enzyme pyruvate kinase catalyses the
transfer of high energy phosphate from PEP
to ADP, leading to the formation of ATP.
This step is also a substrate level
phosphorylation.
Pyruvate kinase
Phosphoenolpyruvate Pyruvate
Mg+2
ADP ATP
Glucose
ATP
Mg+2 HK or GK
ADP
Glucose 6-Phosphate
Mg+2 Phosphohexose isomerase
Fructose 6-Phosphate
ATP
Mg+2 Phosphofructokinase
ADP
Fructose 1, 6-bisphosphate
Aldolase
DHAP Glyceraldehyde 3-Phosphate
Phosphohexose isomerase
DHAP Glyceraldehyde 3-Phosphate
NAD Glyceraldehyde 3P-
Pi dehydrogenase
Iodoacetate,
NADH+H+ Arsenate
1,3-bisphosphoglycerate
ADP
Phosphoglycerate
Mg+2 kinase
ATP
3P-glycerate
Mutase
2P-glycerate
2-Phoglycerate
Enolase
Mg+2
Fluoride
H2O
Phosphoenolpyruvate
ADP
Mg+2 Pyruvatekinase
ATP
Pyruvate
NAD
Lactate
dehydrogenase
NADH+H+
Lactate
Regulation of glycolysis
Three regulatory enzymes:
Hexokinase & glucokinase
Phosphofructokinase
Pyruvate kinase
Catalysing the irreversible reactions
regulate glycolysis.
Hexokinase
Hexokinase is inhibited by glucose 6-
phosphate.
This enzyme prevents the accumulation of
glucose 6-phosphate due to product
inhibition.
Glucokinase
Glucokinase, which specifically
phosphorylates glucose, is an inducible
enzyme.
The substrate glucose, probably through
the involvement of insulin, induces
glucokinase
Phosphofructokinase (PFK)
Phosphofructo kinase (PFK) is the most
important regulatory enzyme in glycolysis
PFK is an allosteric enzyme regulated by
allosteric effectors ATP, citrate & H+ ions (low
pH) are the most important allosteric
inhibitors.
Fructose 2 ,6-bisphosphate, ADP, AMP & Pi are
the allosteric activators.
Role of fructose 2,6-bisphosphate in glycolysis
Fructose-2,6-bisphosphate (F2,6-BP) is
considered to be the most important
regulatory factor (activator) for controlling
PFK & ultimately glycolysis in the liver.
F2,6-BP is synthesized from fructose 6-p by the
enzyme phosphofructokinase called PFK-2
(PFK-1 is the glycolytic enzyme)
F2,6-BP is hydrolysed by fructose 2,6 -
bisphosphatase.
The function of synthesis & degradation of F2,6-BP
is brought out by a single enzyme (same
polypeptide with two active sites) which is
referred to as bifunctional enzyme.
The activity of PFK-2 & fructose 2,6- bisphosphatase
is controlled by covalent modification which, in
turn, is regulated by c AMP.
Cyclic AMP brings about
dephosphorylation of the bifunctional
enzyme, resulting in inactivation of active
site responsible for the synthesis of F2,6-BP
but activation of the active site responsible
for the hydrolysis of F2,6-BP
Pyruvate kinase
PK Inhibited by ATP & activated by F1,6-BP.
Pyruvate kinase is active (a) in
dephosphorylated state & inactive (b) in
phosphorylated state.
Inactivation of pyruvate kinase is brought
about by cAMP-dependent protein kinase.
The hormone glucagon inhibits hepatic
glycolysis by this mechanism.
Energy yield from glycolysis
During anaerobic:
One molecule of glucose is converted to 2
molecules of lactate, there is a net yield of 2
molecules of ATP.
4 molecules of ATP are synthesized by 2
substrate level phosphorylation.
2 ATP molecules are used in steps 1 & 3,
Hence, net yield is 2 ATP.
During Aerobic condition
2 NADH molecules, generated in the
glyceraldehyde 3P-dehydrogenase
reaction & enter ETC.
NADH provides 3 ATP, this reaction
generates 3x2=6 ATP
Total ATP is 6+2=8 ATP.
Conversion of pyruvate to lactate
In anaerobic condition, pyruvate is reduced
to lactate by lactate dehydrogenase (LDH).
LDH has 5 iso-enzymes.
The cardiac iso-enzyme of LDH will be
increased in myocardial infarcts.
Conversion of pyruvate to lactate
Significance of Lactate Production
The NADH is obtained from the reaction
catalysed by glyceraldehyde 3-phosphate
dehydrogenase.
The formation of lactate allows the
regeneration of NAD+ which can be reused by
glyceraldehyde 3-phosphate dehydrogenase.
Glycolysis proceeds even in the absence of
oxygen to supply ATP.
Reconversion of NADH to NAD+ during anaerobiasis
Glycolysis is very essential in skeletal muscle
during strenous exercise where oxygen
supply is very limited.
In RBCs, there are no mitochondria.
Glycolysis in the erythrocytes leads to
lactate production
RBCs derive energy only through glycolysis,
where the end product is lactic acid.
Lactic acidosis
Elevation of lactic acid in the circulation
(normal plasma 4-15 mg/dl) may occur due to
its increased production or decreased
utilization.
Mild forms of lactic acidosis are associated
with strenuous exercise, shock, respiratory
diseases, cancers, low PDH activity, von
Gierke's disease etc.
Severe forms of lactic acidosis are observed
due to impairment/collapse of circulatory
system -in myocardial infarction, pulmonary
embolism, uncontrolled hemorrhage & severe
shock.
This type of lactic acidosis is due to
inadequate supply of O2 to the tissues with a
drastic reduction in ATP synthesis, which may
lead to death.
Oxygen debt refers to the excess amount
of O2 required to recover.
Measurement of plasma lactic acid is
useful to know about the oxygen debt,
and monitor the patient's recovery.
Pasteur effect
The inhibition of glycolysis by oxygen
(aerobic condition) is known as Pasteur
effect.
Pasteur effect is due to the inhibition of the
enzyme phosphofructokinase.
Glycolytic intermediates from fructose 1,6-
bisphosphate onwards decrease while the
earlier intermediates accumulate.
Crabtree effect
Inhibition of oxygen consumption by the
addition of glucose to tissues having high
aerobic glycolysis is known as Crabtree effect.
Opposite to that of Pasteur effect.
Crabtree effect is due to increased competition
of glycolysis for inorganic phosphate (Pi) &
NAD+ which limits their availability for
phosphorylation & oxidation.
References
Textbook of Biochemistry – U Satyanarayana
Harper
Textbook of Biochemistry – DM Vasudevan
Lippincott
Textbook of Biochemistry – MN Chatterjea
