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Glucose Metabolism
Pratt and Cornely, Chapter 13
Glycolysis Expectations
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Memorize/learn Figure 13.2
Know overall reaction and stages
Explain chemical logic of each step
Enzyme mechanisms presented in book
Glycolysis
• Ten enzymes that take
glucose to pyruvate
• Cytosol
• ATP and NADH
Reactions and Enzymes of Glycolysis
ATP
ATP
Pi + NAD+
ADP
2x
ADP
ADP
NADH
ADP
2x
2x
ATP
2x
ATP
• Hexose and triose
phases
• Energy input and
payoff phases
Energy Input
Energy Payoff
Know...
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Substrates
Co-substrates
Products
Enzyme names
1. Hexokinase
• Previous concepts: Induced fit, kinase
• Utilizes 1 ATP
• Chemical logic?
Problem 3
• (Notice miswording) The DGo’ value for
hexokinase is -16.7 kJ/mol, but it is twice as
spontaneous under cellular conditions.
– What is the ratio of G-6-P to glucose under cellular
conditions when the ratio of ATP:ADP is 10:1?
– How high would the ratio of G-6-P to glucose have
to be to reverse the hexokinase reaction by mass
action?
2. Phosphoglucose Isomerase
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Previous concepts: Isomerization
CONCEPT: Near-equilibrium
Chemical logic:
Stereochemistry—reverse does not produce mannose!
3. PFK-1
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Previous concepts: Allosteric inhibition
Utilizes 1 ATP
Pathway logic:
First committed step of glycolysis
– Why?
– regulation
Regulation: Bacteria vs. Human
4. Aldolase
• Previous concepts: Standard free energy is
+23kJ, but it is a near equilibrium reaction
• Pathway logic:
• Beginning of triose stage
Aldolase
Mechanism
5. Triose Phosphate Isomerase
• Previous concepts: Catalytic perfection
• Pathway logic:
• Most similar to which previous reaction?
6. Glyceraldehyde-3-P DH
• Previous concepts: Redox and dehydrogenase
• Pathway logic: Utilizes negative free energy of
_____________ to drive nonspontaneous
formation of ___________________
GAPDH Mechanism
Be able to draw mechanism with full NAD+ structure
7. Phosphoglycerate Kinase
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Previous concepts: High energy bond
Produces 2 ATP: substrate level phosphorylation
Pathway logic?
Coupled to reaction 6 by Le Chatelier
Coupled Reactions
• GAPDH = 6.7 kJ/mol
• PG Kinase = -18.8 kJ/mol
• Overall:
A set of near-equilibrium reactions at cellular concentration
8. Phosphoglycerate Mutase
• Previous concepts: Covalent catalysis
• Pathway logic:
• Mutase—isomerization with P transfer
Mechanism
• Not a simple transfer
• What happens if the bisphosphate escapes?
9. Enolase
• Concept: Phosphoryl group transfer potential
• Chemical logic?
10. Pyruvate Kinase
• Production of 2 ATP
• VERY high energy bond allows formation of
_________ while still being irreversible
• Regulation: F-1,6-BP can act as a feed-forward
activator to ensure fast glycolysis
Overall Energetics
• Standard Free
energies are up and
down
• Free energies under
cellular conditions
are downhill or near
zero
– Three irreversible
Fate of Pyruvate
Amino acid
and nitrogen
metabolism
Gluconeogenesis
Aerobic
Energy
Anaerobic in
higher organisms
Anaerobic in
microorganisms
The Problem of Anaerobic Metabolism
• With oxygen, the NADH produced in glycolysis
is re-oxidized back to NAD+
• NAD+/NADH is a co-substrate which means…
• If there is no oxygen, glycolysis will stop
because…
• The solution to the problem is to…
The solution in Yeast
• Pyruvate is decarboxylated to
acetaldehyde
• Acetaldehyde transformed to
ethanol
– What type of reaction?
– What cofactor?
• NAD+ is regenerated to be
reused in GAPDH
The Solution in Us
• Lactate formation: Draw a mechanism
• Balanced equation
We don’t operate anaerobically...
• Most energy still
trapped in lactate
• Back to pyruvate,
then acetyl-CoA
• Citric acid cycle
Other sugars enter glycolysis
High fructose diet
puts sugars through
glycolysis while
avoiding major
regulation step
Glucose Metabolism
Overview
• Keep the main
pathway
purposes distinct
• But learn details
of chemistry and
regulation based
on similarities
Glucose Metabolism
Overview
• Gluconeogenesis
• Glycogen
metabolism
• Pentose
Phosphate
Pathway
Precursors for Gluconeogenesis
• Names of
compounds?
• Type of reaction?
• Type of enzyme?
• Cofactor(s)?
Chemistry of Gluconeogenesis
• Pyruvate transformed to glucose
• But energetically costly—no perpetual motion
machine!
• Points of regulation
Glycolysis
• Gluconeogenesis
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• Step 10: no ATP
back
• Step 8: no ATP back
• Step 3: costs 2 ATP
• Step 1a/b: costs 4
ATP equivalents
Step 1: costs 1 ATP
Step 3: costs 1 ATP
Step 7: makes 2 ATP
Step 10: makes 2
ATP
Step 1a
• Pyruvate Carboxylase
– Biotin
– Costs ATP to make driving force for next reaction
• TIMES TWO!
– First step in biosynthesis of glucose and many other
molecules
• Related to which amino acid?
Mechanism
• Mixed anhydride
activates carboxyl
group
• Coupled through
biotin coenzyme
• Enolate attacks
activated
carboxylate
Step 1b
• PEP carboxykinase
– ATP cost to restore PEP
• TIMES TWO!
– CO2 loss drives rxn
Step 8
• Fructose-1,6-bisphosphatase
• No additional energy input
• Phosphate ester hydrolysis is spontaneous
Step 10
• Glucose 6-phosphatase
– Liver (and others)
– Not in muscle
Problem 34
• A liver biopsy of a four-year old boy indicated
that the F-1,6-Bpase enzyme activity was 20%
normal. The patient’s blood glucose levels
were normal at the beginning of a fast, but
then decreased suddenly. Pyruvate and
alanine concentrations were also elevated, as
was the glyceraldehyde/DHAP ratio. Explain
the reason for these symptoms.
Key Regulation
• At the committed step in glucogenic cells
• Principle of Reciprocal Regulation
• Local regulation vs Hormone regulation
Key Regulation
• Local regulation
– AMP/ATP (energy charge)
– Citrate (feedback)
• Hormone regulation
– Fructose-2,6-bisphosphate
• Gluconeogenesis is inhibited
• Glycolysis is stimulated
Problem 39
• Brazilin, a compound found in aqueous
extracts of sappan wood, has been used to
treat diabetics in Korea. It increases the
activity of the enzyme that products F-2,6-BP
and stimulates the activity of pyruvate kinase.
What is the effect of adding brazilin to liver
cells in culture? Why would brazilin be an
effective treatment for diabetes?
Glucose Metabolism
Overview
• Gluconeogenesis
• Glycogen
metabolism
• Pentose
Phosphate
Pathway
Glycogen
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Storage molecule
Primer necessary
Very large!
Multiple ends allow
for quick synthesis
and degradation
Chemistry of Synthesis
• Step 1
• Near equilibrium
• The link to glucose-6-phophate, our central
molecule
Chemistry of Synthesis
• Step 2
• Count high energy
bonds
• Pyrophosphatase
– Common motiff
• UDP-glucose:
activated for
incorporation
Chemistry of Synthesis
• Step 3
• Glycogen
synthase
• Growing end is
non-reducing
• UDP released
Branching
Energetics of Synthesis
• Total cost: one ATP equivalent from G-6-P
Four Degradation Enzymes
• Two enzyme debranching process
• Glycogen phosphorylase
• phophoglucomutase
Chemistry of Degradation
• Glycogen
phosphorylase
• Key Regulation site
• Inorganic phosphate
as a nucleophile
• Remake G-1-P with
no ATP cost
Debranching
• Transfer branch
• a(16)glycosidase
Overall Energetics
Key Regulatory Enzymes
Glycogen Storage Diseases
Many disrupt glycogen breakdown in muscle and/or liver
(hypoglycemia, enlarged liver, muscle cramps...)
Glucose Metabolism
Overview
• Gluconeogenesis
• Glycogen
metabolism
• Pentose
Phosphate
Pathway
Pentose Phosphate Pathway
• Dual Purpose
– Synthesis of “reducing potential”
– Synthesis of 5-carbon sugars
• At cost of one carbon worth of carbohydrate
• Net reaction:
Physiological Purposes
Glutathione:
Protection from Oxidation
• Glutathione is the redox
buffer of the cell
• Regenerated by NADPH
• PPP especially
important in RBC
because it is only means
to generate reducing
power (no
mitochondria)
Complex, 2-Stage Process
• Oxidative Stage
– Generates
reducing power
and ribose
• Non-oxidative
stage
– Regenerates 3- and
6-carbon sugars
from 5 carbon
sugars
Oxidative Stage Step 1:
• G-6-P DH
• Lactone formation
Oxidative Stage Step 2:
• Also a spontaneous hydrolysis
• Practice mechanism, carbohydrate orientation
Oxidative Stage Step 3:
• Oxidative decarboxylation
– Which is first, oxidation or decarboxylation?
• We will see this process again
Biosynthesis of Ribose
Non-oxidative Stage
• To understand purpose, realize that we
generally need to make much more NADPH
than ribose
• Problem: stuck with C5, but need C6 and C3
• Solution: “Shunt” C5 back to C6 through
near-equilibrium reactions
PPP Reactions
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Epimerase
Isomerase
Transketolase
Transaldolase
Epimerase
Transketolase
• Use cofactor (B1) to overcome chemical problem
Mechanism
Transketalase, Transaldolase
• Transketalase
– Transfer of an acyl anion
– Requires TPP to stabilize
unstable anion
• Transaldolase
– Transfer of somewhat
stable enolate ion
– Schiff base helps
• Be able to recognize
need for TK vs. TA
Transketalase or transaldolase?
Different Modes for Different Purposes
Problem 58
• A given metabolite may follow more than one
metabolic pathway. List all possible fates of
glucose-6-P in (a) a liver cell and (b) a muscle
cell.
Summary
of glucose
metabolism