The pay-off phase

• Converts glyceraldehyde-3-P to pyruvate

• Generates 4 ATP, 2 ATP/pyruvate

Remember each of these reactions is run in duplicate for 1 molecule of glucose (each glucose produces 2

glyceraldehyde 3-P)

Rxn 6: Glyceraldehyde-3-P to 1,3-bisphoshoglycerate

C. i. Glyceraldehyde-3-phosphate

dehydrogenase

ii. Dehydrogenase

iii. Also called an oxidoreductase

D. Energy producing step! This

Chemistry C483 Fall 2009 Prof Jill Paterson 30-1

B. Mechanism

D. Energy producing step! This

step produces NADH

E. Not a regulatory step

F. Energy:

Rxn ATP

used

ATP

made

NAD

used

NADH

made

This step

Overall

Rxn 7: 1,3-bisphosphoglcerate to 3-phosphoglycerate

B. Mechanism is reverse of phosphorylation

C. i. Phosphoglycerate kinase

ii. Transferase- kinase! (same as rxn 1 & 3)

D. Purpose: This is first step where ATP is

made!

E. Not a regulatory step

F. Energy

Rxn ATP

used

ATP

made

NAD

used

NADH

made

This step

Overall

Rxn 8: 3-phosphoglycerate to 2-phosphoglycerate

C. i. Phosphoglycerate mutase

ii. Isomerase

iii. Mutase (same as 2 & 5, mutase is enzyme that transfers

phosphoryl group)

D. Purpose: Getting ready for the next enzyme

E. Not a regulatory step

F. Energy

Rxn ATP

used

ATP

made

NAD

used

NADH

made

This step

Overall

Chemistry C483 Fall 2009 Prof Jill Paterson 30-2

Rxn 9: 2-phosphoglycerate to phosphoenolpyruvateB. Mechanism

C. i. Enolase

ii. Lyase

iii. non-hydrolytic cleavage

D. Purpose: To generate a high energy bond

PEP has a higher phosphoryl group transfer potential than 2-PG

E. Not a regulatory step

F. EnergyF. EnergyRxn ATP

used

ATP

made

NAD

used

NADH

made

This step

Overall

Rxn 10: PEP to pyruvateB. Mechanism is same as previous

C. i. Pyruvate kinase

ii. Transferase reaction (kinase)

D. Purpose: ATP is made!! More energy!!

E. This is a regulated step!! Not reversible

∆∆∆∆G = -31.7 kJ/mol

F: Energy

Rxn ATP

used

ATP

made

NAD

used

NADH

made

This step

Overall

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Investment v. payoff balance sheet

Rxn ATP change/glucose NADH change/glucose

1

3

6

7

10

Total

Thus our payoff is ____________________

What happens to pyruvate?

Pyruvate can take 1 of 4 paths next:

Aerobic conditions

1. Converts to acetyl CoA (by pyruvate dehydrogenase) for use in the TCA cycle and

oxidative phosphorylation (leads to more ATP production)

2. Converts to oxaloacetate , which can then shuttle into the synthesize of glucose 2. Converts to oxaloacetate , which can then shuttle into the synthesize of glucose

(gluconeogenesis)

Anaerobic conditions

3. It is converted to Lactate (animal muscles; lactate used in gluconeogenesis)

4. It is converted to ethanol (yeast; alcohol fermantation)

Chemistry C483 Fall 2009 Prof Jill Paterson 30-4

What happens to the NADH?

Need to regenerate NAD+.

Aerobic conditions:

Electron transport system (lectures 33 & 34)

Anaerobic conditions:

EtOH production consumes

Lactate production consumes

Regulation of glycolysis

Our cells must ALWAYS have energy, so glycolysis is HIGHLY regulated

As ATP and NADH levels go up and down, glycolysis goes up and down due to specific regulation

We listed:

7 steps as unregulated (2, 4-9)

3 steps (1, 3, 10) as regulated.

Why are these three steps the regulatory steps?

Steps 2, 4-9:

• These are the unregulated steps

• Have ∆G close to zero

• Essentially at equilibrium

• Can run in either direction

• (And they are common steps in glucose

synthesis (gluconeogenesis))

Steps 1, 3, 10:

• These are the regulated steps

• Have large ∆G

• Therefore not at equilibrium

Chemistry C483 Fall 2009 Prof Jill Paterson 30-5

Regulation at step 1: hexokinase

• Regulates the entry of glucose into glycolysis by controlling the amount of

glucose-6-P.

• This reaction is controlled by FEEDBACK inhibition

• Hexokinase is inhibited by glucose-6-P (its product!):

If too much product, inhibits the production of more by turning

off hexokinase.

Regulation at Step 3: phosphofructokinase-1 (PFK-1)

• This is the major control point of glycolysis!

-Once this step happens, we MUST form pyruvate

-∆G is too large to overcome (-16.7 kJ/mol)

-no alternative pathways

• Multiple things regulate PFK-1

-ATP

-AMP

-Citrate

-Fructose-2,6-bisphosphate-Fructose-2,6-bisphosphate

• A substrate

(active site has a high affinity for ATP)

• An allosteric inhibitor

(inhibition site has a lower affinity for ATP)

Therefore, when ATP concentration low, will only bind at active site. When ATP

concentration is high, it will bind at both sites.

• If [ATP] is high, PFK-1 is turned off.

• If [ATP} is low, PFK-1 is turned on.

This should seem logical- if we have energy (high ATP) we do not need to make more

If we need more energy (low ATP) we need to make more

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• An allosteric activator

has a low affinity for AMP

• If [AMP] is high, indicates [ATP] is low, so we need more energy!

• If [AMP] is high, will bind PFK-1 and activate it! Thus we will increase pyruvate

(and ATP!) production

Regulation at Step 3: by AMP

Chemistry C483 Fall 2009 Prof Jill Paterson 30-7

Regulation at Step 3: by Citrate

• An allosteric (“feedback”) inhibitor

PFK-1 has a low affinity for citrate

• Citrate is synthesized in the TCA cycle from pyruvate

• Therefore, if pyruvate production is high, citrate production will be high.

• So, when [citrate] high, indicates we do not need more pyruvate, so citrate turns off PFK-1 to

stop glycolysis.

Regulation at Step 3: by Fructose-2,6-bisphosphate

• Fructose-2,6-BP is an allosteric activator

• PFK-2 has a lower affinity for Fructose-6-P

than PFK-1.

• Therefore, at high concentrations of

Fructose-6-P, some will be converted to

Fructose-2,6-bisphosphate.

• If [Fructose-2,6-bisP] gets too high, it will

bind to PFK-1, activating it, and increasing

glycolysis.

Chemistry C483 Fall 2009 Prof Jill Paterson 30-8

Regulation at Step 10: Pyruvate kinase

• High [Fructose 1,6-bisphosphate] activates pyruvate kinase

• High [ATP] inhibits pyruvate kinase

• ATP is both a substrate and an inhibitor of this enzyme!

• Why would we want to regulate this step??

The intermediates between _____ can be shuttled to other

pathways, so their production is not a waste.

Regulation of glycolysis

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How do other carbohydrates enter glycolysis?

1. Starch (glucose polymer)

• In mouth, amylases break starch to glucose monomers

• In stomach, acid breaks starch to glucose monomers

• Glucose is then absorbed through intestinal wall to blood and transported

• Glucose can then enter cells and start glycolysis.

1/3 of glucose goes to the heart and skeletal muscles

1/3 of glucose goes to your brain!

1/3 of glucose to the liver for storage

2. Disaccharides

a. Maltose

b. Sucrose

c. Lactose

mannose – enters from glycoproteins

glycerol- results from the breakdown of fat

Entrance of fructose into glycolysis

• Can enter through liver or muscle

• Liver

-3 additional reactions

- Enters glycolysis as 2 molecules of glyceraldehyde-3-P

Chemistry C483 Fall 2009 Prof Jill Paterson 30-10

•Muscle

-Enters the pathway as fructose-6-P

-This is just 1 additional step!!

-We use 1 ATP to get to this entry point

-A muscle specific kinase can phosphorylate fructose

How much energy do we get from fructose?

But we miss at least 1 regulatory step!

• We miss step 1 (in both cases), and step 3 (in liver)

• This takes away regulatory steps.

• People who take in large amounts of fructose tend to have additional fat in their livers.• People who take in large amounts of fructose tend to have additional fat in their livers.

-There is an overproduction of pyruvate

- Pyruvate leads to the production of fats and cholesterol

How does galactose enter glycolysis?

• Galactose requires 5 reactions to transform it to glucose-6-phosphate

On your own:

1. Determine where galactose enters glycolysis

2. Do an energy count- do we get the same amount of energy for galactose?

3. Is regulation different from glucose?Chemistry C483 Fall 2009 Prof Jill Paterson 30-11

Mannose

On your own:

1. Determine where mannose enters glycolysis

2. Do an energy count- do we get the same amount of energy for mannose?

3. Is regulation different from glucose?

Glycerol

• Glycerol is released during the degradation of fatty acids

• 2 reactions to become dihydroxyacetone

-Phosphoryl transfer

-Oxidation

-enters glycolysis as dihydroxyacetone

On your own:On your own:

1. Determine where glycerol enters glycolysis

2. Do an energy count- do we get the same amount of energy for glycerol?

3. Is regulation different from glucose?

What you need to know for all metabolic reactions:

We have discussed glycolysis specifically. For all metabolic reactions we discuss in lecture, you

should be able to:

Materials Flow: Trace the fate of labeled carbon or other elements through the pathway.

Energy Flow: Trace the production and consumption of "high energy" compounds like ATP.

Electron Flow: Trace the production and consumption of reducing power.

Net Flows: Compute a) free-energy changes, b) net gain or loss of ATP, and c) net redox change

during all of any segment of the pathway.

Regulation: Explain the effects of allosteric effectors, hormones, second messengers, and reversible

phosphorylation on individual enzymes and on overall flow through the pathway.

Connections: Describe connections to other pathways.

Push electrons for only the presented mechanisms.

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Metabolic pathways of C483

Chemistry C483 Fall 2009 Prof Jill Paterson 30-13