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Glycolysis, gluconeogenesis and the TCA cycle
Prof. Alison Baker
BLGY1115 L9
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TCA cycle
Glucose
Pyruvate
Acetyl CoA
Fatty acids
citrateAcetyl CoA
GlycolysisGluconeogenesis
Fatty acid synthesis
Fatty acid
degradation
NADH
FADH2
ATP
Overview of Metabolism
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Glycolysis-Takes place in the cytoplasm
Note –
which reactions require ATP
which synthesise ATP
Which synthesise NADH
many reactions are reversible
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Substrate level phosphorylation
A phosphate group is transferred from one compound to another withoutthe involvement of electron transfer reactions as occur when ATP is synthesised in mitochondria as we shall see later.
All that is required is that the standard free energy of hydrolysis Go’of the compound donating the phosphate group is larger and more negative
than the Go’ of hydrolysis of the phosphate containing compound that is formed.
1,3 bisphosphoglycerate → 3-phosphoglycerate Go’ = -49.3 kJ mol-1
ADP + Pi =H+→ ATP + H2O Go’ = + 30.5 kJ mol-1
The enzyme phosphoglycerate kinase couples these reactions allowing the large negative G of the first reaction to be used to drive the energetically unfavourable second reaction.
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Figure 2-72a Molecular Biology of the Cell (© Garland Science 2008)
Phosphoglyceratekinase
Glyceraldehyde 3 Phosphate dehydrogenase
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Figure 2-73 Molecular Biology of the Cell (© Garland Science 2008)
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Glycolysis net reaction
Glucose +2 ADP + 2Pi + 2NAD+ →2 pyruvate + 2ATP+ 2NADH
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Quick self test
find the WRONG answer
1. Glycolysis takes place in the cytoplasm
2. Uses 2 ATP per glucose at the beginning of
the pathway
3. Splits a 6 C sugar phosphate to two 3C sugar
phosphates
4. Yields 2 molecules of NADH per glucose
5. Yields 2 molecules of ATP by substrate level
phosphorylation
6. Overall yield is 2 ATP per glucose
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Glycolysis net reaction
Glucose +2 ADP + 2Pi + 2NAD+ →2 pyruvate + 2ATP+ 2NADH
Q. What happens to the pyruvate and NADH?
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Under anaerobic conditions NAD+ is regenerated
2 pyruvate 2 lactate
2NADH 2NAD+
e.g in muscle
2 pyruvate 2 ethanol + 2 CO2
2NADH 2NAD+
e.g alcoholic fermentation in yeast
More on fermentation from Prof Denecke later
This is essential for anaerobic glycolysis to be sustained
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Cori cycle
Glucose
Pyruvate
Lactate
2ATP
2 NAD+
Muscle
Anaerobic glycolysis
blood
Glucose
Pyruvate
Lactate
4ATP
NADH
2 NADH
Liver
Gluconeogenesis
2 GTP
Lactate produced by anaerobic glycolysis in the sprinter’s musclesis transported to the liver and reconverted to glucose.
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Gluconeogenesis is not simply the reverse of glycolysis
ADP + Pi + PEP → pyruvate + ATP Pyruvate kinase (glycolysis)
Pyruvate + ATP + CO2 → oxaloacetate + ADP + Pipyruvate carboxylase (gluconeogenesis)
Oxaloacetate + GTP → PEP + GDP + CO2PEP carboxykinase (gluconeogenesis)
The pyruvate kinase reaction is essentially irreversible (Go’ -62.8 kJ mol-1) so 2
additional reactions using high energy phosphate compounds are required to bypass
this step.
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Gluconeogenesis proceeds by the reverse ofGlycolysis as far as fructose 1,6 bisphosphate
Two different enzymes,fructose 1,6 bisphosphatase andglucose 6 phosphatase are required to reverse the twoATP requiring steps of glycolysis catalysed by Hexokinase and phosphofructokinase
These enzymes are heavily regulated to avoid glycolysisand gluconeogenesis both being active at the same time
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Gluconeogenesis
Note –
use of 2 high energy phosphate bonds to bypass pyruvate kinase
distinct enzymes to reverse the two ATP requiring steps of glycolysis
oxaloacetate
ATP + CO2ADP + Pi
GTP
GDP + Pi + CO2
Glucose-6 phoaphatase
Fructose 1,6bisphosphatase
Pyruvate carboxylase
PEP carboxykinase
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Quick self test 2
find the WRONG answer
1. When there is insufficient oxygen pyruvate is converted to lactate.
2. Lactate formation allows oxidation of NADH so that glycolysis can continue
3. Lactate can be converted back to glucose in the liver
4. The pathway that does this is called gluconeogenesis
5. Gluconeogenesis is the exact reverse of glycolysis
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Glycolysis net reaction
Glucose +2 ADP + 2Pi + 2NAD+ →2 pyruvate + 2ATP+ 2NADH
Q. What happens to the pyruvate and NADH?
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Under aerobic conditions pyruvate and NADH from glycolysis are oxidised in mitochondria
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Site of TCA cycle
Site of electron transfer reactions and ATP
Synthesis. Highly impermeable.
Permeable to molecules of up to 1000 Da
The impermeability of the inner membrane is essential for ATP synthesis, but means
that transporter proteins are required to move metabolites in and out of mitochondria
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The inner membrane contains specific transport proteins forimportant metabolites
We shall return to how transport is energised in the next lecture.
20The malate-aspartate shuttle for transporting reducing equivalents
Malate dehydrogenase
Aspartate amino transferase
Aspartate amino transferase
Malate dehydrogenase
NAD+ and NADH do not have a transport protein butneed to cross the inner membrane
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Quick self test 3
The mitochondrial inner membrane is highly
impermeable and specific transporters are
required to move metabolites between the
matrix and the cytosol.
True or False?
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Pyruvate is transported into the mitochondrial matrix and converted to Acetyl CoA by Pyruvate Dehydogenase
This step is an important site of regulation of the TCA cycle (see later)
Pyruvate + NAD+ + CoA→ acetyl CoA + CO2 + NADH
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Overview of the TCA cycle
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The TCAcycle
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Can you work out what the net reaction of the cycle is?
Acetyl CoA + 3NAD+ +FAD + GDP + Pi +2H2O →CoASH +3(NADH +H+) +FADH2 +GTP +2CO2
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The TCA cycle is a source of biosynthetic precursors
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‘Topping up’ the cycle
If intermediates that are removed for biosynthesis are not replenished the cycle
will stop
Note that all carbon that enters as the acetyl group is oxidised to CO2, so
Simply putting in more acetyl CoA is no help
Pyruvate carboxylase provides a means of replenishing oxaloacetate
Pyruvate + CO2+ATP+H2O → Oxaolacetate + ADP +Pi + 2H+
We have met this reaction before in gluconeogenesis
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Control of the cycle
Pyruvate dehydrogenase
a-ketoglutarate
dehydrogenase
Succinyl CoA
synthetase
The products of the cycle,
ATP and NADH
inhibit it by activating
a protein kinase that
phosphorylates and inhibits
PDH
There is also regulation
of PDH activity by
hormones, e.g.
Insulin stimulates
PDH dephosphorylation
and therefore activation
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Quick self test 4
find the INCORRECT answer
1. The TCA cycle completely oxidises acetyl
groups supplied as acetyl CoA
2. The products of the TCA cycle are CO2, GTP,
NADH and FADH2 and CoA
3. Intermediates can be removed from the cycle
for biosynthetic reactions.
4. Pyruvate dehydrogenase converts pyruvate to
oxaloacetate allowing the intermediates of the
cycle to be topped up.
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Reading
• You will find the material covered in these 3 lectures in any good biochemistry text book.
• Some examples from the libraryEssential biochemistry / Charlotte W. Pratt, Kathleen Cornely.
• ed. Hoboken, N.J. : Wiley ; Chichester : John Wiley [distributor], c2011. c2011
• Biochemistry / Mary K. Campbell, Shawn O. Farrell.
•6th ed. : international student ed.
Pacific Grove, Calif. : Brooks/Cole ; London : Thomson Learning [distributor], c2009. c2009
• Principles of biochemistry / Donald Voet, Judith G. Voet, Charlotte W. Pratt.
•3rd ed., International student version.
Hoboken, N.J. : Wiley, c2008. c2008