Biochemistry
Topic 9: Glycolysis – steps, energy generation, regulation, and interrelationships to other pathways
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The glycolysis pathway consists of 10 steps which can be divided into 2 groups the
preparatory phase and the payoff phase
Steps 1, 3 and 10 are regulated. Step 3 is highly regulated since there is “no turning back”
from it.
Glycolysis can function either aerobically or anaerobically
Glycolysis is both the principal route for glucose metabolism and the main pathway for the
metabolism of fructose, galactose, and other carbohydrates derived from the diet.
The Steps of glycolysis:
The preparatory phase:
Step
Substrate
1
Glucose
Glucose-6phosphate
(Glu-6-P)
Fructose-6phosphate
2
3
Fructose-1,6biphosphate
4
5
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Product
Glucose-6phosphate
Fructose-6phosphate (Fru-6P)
Fructose-1,6biphosphate
Dehydroxyacetonephosphate and
glycer-aldehyd-3phosphate
DehydroxyGlycer-aldehyd-3actetonephosphate
phosphate
The payoff phase:
Enzyme
Hexokinase /
Glucokinase
Phosphofructokinase
1 (PFK1)
6
Gylcer-aldehyd-3phosphate
1,3bisphosphoglycerate
7
1,3bisphosphoglycerate
3-phosphoglycerate
8
3-phosphoglycerate
2-phosphoglycerate
Keto-enol
tautomery can
also happen
ATP ADP
Irreversible step
Enzyme
Glyceraldehyde3-phosphate
dehydrogenase
(GAP-DH)
Phosphoglycerat
e kinase
Phosphoglycerat
e mutase
In
Out
NAD
NAD
H
+
ADP
Using
inorganic
phosphat
e
ATP
Phosphoenolpyruvat
Enolase
e
Phosphoenolpyruvat
Pyruvate (first Enol
Pyruvate kinase
ADP
ATP
e
then Keto)
In total: Glucose + 2NAD+ + 2ADP + 2Pi  2 pyruvate + 2NADH + 2ATP + 2H2O
The total energy stored in glucose is: -2840kJ/mol. Glycolysis only generates about 5.2% of
this energy which is -146kJ/mol
Glycolysis is controlled by the anount of ATP in a cell the more ATP there is in a cell the less
glycolysis there is in the cell
2-phophoglycerate
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Phosphorilation
Phosphotrioseisomerase
Product
-
ATP ADP
Aldolase enzyme
Substrate
10
Out
Phosphohexoseisomerase
Ste
p
9
In
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Some substances can inhibit the glycolysis enzymes:
Blocked enzyme:
Enolase
Glyceraldehyde-3phosphate
dehydrogenase
GAP-DH
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-
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Blocked by:
Fluouride
What happens?
2-phosphoglycerate cannot become
phosphoenol pyruvate
What for?
To estimated blood
glucose levels and
the prevention of
glycolysis of the cells
in the blood
Iodoacetate No further phosphorilation of the
glyceraldehyde-3-phosphate and
therefore the 1,3,bisphophoglycerate
will not be formed. The glycolysis will
stop at this stage.
There are 3 main regulations in the glycolysis itself:
o Step 1 – phosphorilation of glucose. The enzyme, hexokinase, is inhibitied by its
products: Feedback inhibition. If there is too much glucose-6-phosphate, it will
inhibit the enzyme to produce more glucose-6-phosphate. If the gluscose is not
phosphorylated it can diffuse back to the blood and supply other cells that might
need it. The main regulator of the blood’s glucose is the liver. Extra G6P is converted
into G1P and will be stored as glycogen. In the liver, however, the enzyme
glucokinase is active. Glucokinase is not inhibited by G6P. This allows a continous
production of fatty acids, cholesterol, and glycogen in the liver, even if the
hexokinase activity is low.
o Step 3 – phosphorilation of fructose-6-phosphate. The PFK1 enzyme is tightly
regulated because this step is irreversible. The PFK1 enzyme’s activity will be
enhanced when the ADP and AMP levels will rise in the cell or the AP levels are too
low. Fructose-2,6-biphosphate (from PFK2) is a strong allosteric activator for the
PFK1. Large amounts of other fuel substances, like fatty acids, in the cell will also
inhibit the function of PFK1
o Step 10 – dephosphorylation of the phosphoenolpyruvate to form pyruvate. All of
the pyruvate kinase isoenzymes are allosterically inhibited by ATP, Acetyl-CoA and
long fatty acids. The liver pyruvate kinase can be further inhibited by phosphorilation
– it is phosphorylated by cAMP-dependet protein kinase.
o These 3 steps are characterized by large negative G values. In order to reverse
these steps it would be necessary to overcome this high energy barrier. Because of
this high energy barrier these reactions are irreversible in most cells.
During starvation fatty acids are being oxidized, there will be a decrease in the activity of
glycolysis. This is because the important steps are regulated not only by their products and
ATP, but also by fatty acids in the cell.
The oxidation of pyruvate to acetyl-CoA is irreversible and will send the pyruvate to the citric
acid cycle. Inhibiting the pyruvate dehydrogenase, can lead to lactic acidosis. This will happen
due to the lack of ATP, which causes the pyruvate to be sent to lactic acid fermentation. The
excess of lactic acid will result in acidosis.
The aspect of pyruvate after the glycolysis:
o Usually pyruvate will be sent to the mitochondria for the citric acid cycle. There the
remaining energy of the glucose will be obtained. The process requires O2 and will
generate 4 CO2, 4H2O, and 36-38 (according to Harper) or 30-32 (according to
Lehninger) ATP molecules. The difference in the ATP can be explained because the
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amount of ATP that can be produced from NADH and FADH2 is different – 2.5 and 1.5
or 3 and 2.
o In absence of O2 (anaerobic conditions), pyruvate can undergo 2 pathways:
 Fermentation to lactate – this happens in muscles which work hard but do
not have enough supply of O2. In this pathway the lactate dehydrogenase
enzyme is active.
 Fermentation to ethanol – this happens in yeast. This will produce 2
molecules of ethanol and 2 molecules of CO2. Pyruvate Decarboxylase and
alcohol dehydrogenase will catalyze these reactions.
Glycolysis can accept substances from many other pathways:
Substance
Pathway:
Where the substance
enters the glycolysis:
Glucose-6-phosphate
Glycogen breakdown
2nd step: Glu-6-P to Fru6-P
Glucose
Starch breakdown
1st step: Glucose to
Glu-6-P
Fructose-6-phosphate
Mannose: This 6 carbon
3rd step: Fru-6-P to
monosaccharide is
Fructose-6phosphorylated by
biphosphate
hexokinase to mannose-6phosphate. This will then be
converted by
mannosephosphate
isomerase to Fru-6-P.
Fructose (as 2 molecules of Fructose is converted by
5th step: dehydroxyglyceraldehyd-3-phoshate)
fructokinase to Fru-1-P,
acetone-phsophate to
which then is converted by
Glycer-aldehyd-3Fru-1-P Aldolase into
phosphate or
glyceraldehyd and
6th step: Gylceraldehyddehydroxy-acetone3-phosphate to 1,3phosphate. Glyceraldehyd
biphospho-glycerate
will be phosphorilized by
glyceraldehyd kinase.
Galactose (as Glu-6-P)
In the liver galactose is
2nd step: Glucose-6converted by galactokinase
phosphate to Fructoseinto Gal-1-P, which is then
6-phosphate.
converted by Gal-1-P uridyl
transferase into Glu-1-P,
which is converted by
phosphoglucomutase to
Glu-6-P.
Oxaloacetate (as PEP)
Oxaloacetate is converted
10th step: pyruvate
by PEP carbokinase to PEP
and then into Pyruvate
Intermediates of the glycolysis can enter some other pathways:
o Glucose-6-phosphate can enter the pentose-phosphate pathway. Glycer-aldehyd-3phosphate from the ppp can enter glycolysis.