Regulation of Glycolytic pathway and TCA cycle

1. Regulation of
Glycolysis &
TCA cycle

2. Hello!
I am Sameer Turki
This presentation has been split into two
parts for better understanding.

3. 1.
Regulation of
Glycolysis

4. REGULATION OF GLYCOLYSIS
Flux through a metabolic pathway can be regulated in
several ways:
1. Availability of substrate
2. Concentration of enzymes responsible for rate-
limiting steps
3. Allosteric regulation of enzymes
4. Covalent modification of enzymes

5.  Three reactions of the glycolysis are the regulatory steps.
1) Conversion of glucose to glucose-6-phosphate catalyzed by
hexokinase.
2) Fructose to fructose1, 6-bisphosphate catalyzed by
phosphofructokinase
3) Formation of pyruvate from PEP catalyzed by pyruvate
kinase.
 Of the 10 steps in the glycolytic pathway, three involve large
negative ∆G and are essentially irreversible. These are steps
1 (phosphorylation of glucose), 3 (phosphorylation of
fructose-6-phosphate) and 10 (transfer of phosphate from
phosphoenolpyruvate to ADP). Net ∆G for glycolysis is about
-23 kcal/mol.
REGULATION OF GLYCOLYSIS

6. In practice, we generally consider reactions where ∆G is
larger than about -2 kcal/mol to be “irreversible”.
REGULATION OF GLYCOLYSIS

7.  The concentration of these three enzymes in the cell is
regulated by hormones that affect their rates of
transcription. Insulin is a peptide hormone secreted by
pancreatic β-cells in response to sudden increases in blood
glucose levels. The general effect of insulin is to promote
the storage of energy when food is available in abundance.
 Glucagon is a different peptide hormone secreted by the
pancreatic α-cells. Its secretion is stimulated by low blood
glucose levels, and its general effect is to oppose the action
of insulin.
 Insulin upregulates the transcription of glucokinase,
phosphofructokinase, and pyruvate kinase, while glucagon
downregulates their transcription.
REGULATION OF GLYCOLYSIS

8. Hexokinase
Hexokinase performs step 1 of glycolysis in most tissues,
including muscle and brain. It has a low Km (high affinity) for
glucose, so it permits initiation of glycolysis even when blood
glucose levels are relatively low. However, its Vmax is relatively
low. Hexokinase is inhibited by the product of its reaction,
glucose-6-phosphate. This is a very important regulatory step,
since it prevents the consumption of too much cellular ATP to
form G6P when glucose is not limiting.
REGULATION OF GLYCOLYSIS

9. Phosphofructokinase
 PFK catalyzes the rate-limiting step in glycolysis and is the
most important control point. It is also the first irreversible
step that is unique to the glycolytic pathway; G6P can be
used as an intermediate in other pathways including
glycogen synthesis and the pentose phosphate pathway.
 PFK is allosterically inhibited by ATP, so glycolysis is slowed
when cellular ATP concentrations are high. ATP binds to a
site on PFK distinct from the active site, causing a
conformational change resulting in rotation of the positions
of Arg162 and Glu161. In the high-affinity state, the positive
charge on Arg162 stabilizes the negative charge on the
phosphate of F6P, and Km is low. In the low-affinity state, the
negative charge on Glu161 repels F6P.
REGULATION OF GLYCOLYSIS

10. The conformational transition between
these two states is also regulated by
cellular pH. Excess H+ ions favor the low
affinity state. Thus when cellular lactate
is high (usually when oxidative
phosphorylation is inhibited), the rate of
glycolysis is reduced, preventing further
accumulation of intracellular acid. This
regulation helps to minimize the risk of
lactic acidosis when oxygen is scarce.
REGULATION OF GLYCOLYSIS

11.  When cellular energy is limited, glycolysis should be
upregulated. PFK is allosterically activated by high levels of
AMP. AMP overcomes the inhibitory effect of ATP.
 Another allosteric activator of PFK is fructose 2, 6
bisphosphate. F-2,6-BP is not an intermediate in the
glycolytic pathway. F-2,6-BP also overcomes the inhibitory
effect of ATP. F-2,6-BP is made from F6P by a specific kinase,
phosphofructokinase 2 (PFK2). F-2,6-BP is also an important
regulator of the process of gluconeogenesis, where glucose is
synthesized from pyruvate.
REGULATION OF GLYCOLYSIS

12. Pyruvate kinase
 Pyruvate kinase is the third regulated enzyme of glycolysis.
Like PFK, pyruvate kinase is regulated both by allosteric
effectors and by covalent modification (phosphorylation).
Pyruvate kinase is activated by F-1,6-BP in the liver, a second
example of feedforward stimulation. ATP and alanine (a
biosynthetic product of pyruvate) act as allosteric inhibitors
of pyruvate kinase.
 Phosphorylation of pyruvate kinase is regulated by blood
glucose level, just like PFK. High glucagon (low blood sugar)
causes phosphorylation, which in this case renders the
enzyme inactive.
REGULATION OF GLYCOLYSIS

13. REGULATION OF GLYCOLYSIS

14. REGULATION OF GLYCOLYSIS

15. Regulation of
TCA cycle
2.

16. Factors that regulate TCA cycle:
i) Substrate availability
ii) Product accumulation
iii) Ratio of NADH/NAD+ and ATP/ADP
 One of the important roles of the cycle is to provide reduced
cofactors, such as NADH and FADH2.
 These reduced cofactors are further oxidized by the electron
transport chain which is localized in the inner mitochondrial
membrane. Energy of the oxidation is conserved in the form of
ATP.
 So, the two most important molecules, which regulate the
TCA cycle, are the ratio of NADH/NAD+ and ATP/ADP. If a cell is
actively metabolizing, which means consuming ATP, ratio of
ATP/ADP will be low, and the ratio of NADH/NAD+ will also be
low.
REGULATION OF TCA CYCLE

17.  It gives a signal to the cell to produce more NADH and ATP to
meet the demand, so the TCA cycle will operate more
efficiently, while in a resting cell, ATP and NADH will
accumulate resulting in high ratios of ATP/ADP and
NADH/NAD+. This will result in inhibition of the activity of the
enzymes responsible for producing them.
REGULATION OF TCA CYCLE

18. Regulation points of TCA cycle
1. The cycle is regulated at the entry level of acetyl-CoA.
Citrate synthase catalyzes the condensation reaction of
acetyl-CoA with OAA. Availability of these substrates will
regulate the activity of citrate synthase, which varies with
the metabolic status of the cell. Accumulation of citrate,
succinyl-CoA and ATP inhibits the activity of citrate
synthase. This inhibition of citrate synthase by ATP is
relieved by ADP accumulation.
2. Second point of TCA regulation is the reaction catalyzed by
isocitrate dehydrogenase. NADH and ATP accumulation
inhibits the activity of the enzyme
.
REGULATION OF TCA CYCLE

19. 3. The third point of
regulation of TCA cycle
is the reaction catalyzed
by the enzyme α–
ketoglutarate
dehydrogenase. Activity
of this enzyme is
inhibited by the
products of the reaction
i.e., succinyl-CoA, and
NADH. When the
products of the reaction
accumulate the activity
of enzyme is inhibited.
REGULATION OF TCA CYCLE

20. REFERENCES:
• Stanford Medicine
http://imed.stanford.edu/curriculum/session6/content/09-
Regulation_of_glycolysis.pdf
• ResearchGate
https://www.researchgate.net/file.PostFileLoader.html?id=577bfd1d40
485412a95d1903&assetKey=AS%3A380558044614656%401467743516991
• Virtual Learning Environment
http://vle.du.ac.in/mod/resource/view.php?id=12106
• Nelson, Cox, (2004), Lehninger Principles of
Biochemistry,4th Edition

21. Thanks!