1. Regulation of Glycolysis/Gluconeogenesis
Citric Acid Cycle / TCA cycle / Kreb’s cycle
Electron Transport Chain (Oxidative Phosphorylation)

2. 3 major points of regulation Phosphofructokinase-1
of glycolysis
Hexokinase
Phosphofructokinase-1
Pyruvate kinase
-32.9
-24.5
Each regulatory reaction
is held far from equilibrium
(large – DG)
Each regulatory step in glycolysis
is coordinately regulated with
gluconeogenesis
Substrate Cycle * *
-26.4

3. Futile/Substrate cycle:
Each regulatory step in glycolysis is coordinately regulated with gluconeogenesis
Futile/Substrate cycle: Pi
ATP
Glycolysis
ADP
H2O
Gluconeogenesis
Sum: ATP + H20  ADP + Pi
 If allowed to occur at the same time – utilization of ATP without useful metabolic work being done
 Cycles provide important means of regulation *

4. Hexokinase Muscle – High affinity for glucose
Usually saturated and working at
maximal rate
Inhibited by glucose 6-phosphate
(product inhibition)
Liver – much lower affinity (higher Km)
for glucose.
Allows liver to process high levels of
glucose
Not inhibited by glucose 6-phosphate
Muscle
Liver

5. Coordinate Regulation of PFK-1 and FBPase-1
(regulation by energy state of cell)
* Commitment step
of glucose into
glycolysis
PFK-1 – inhibited by signals of adequate
energy supplies
ATP
Citrate
Activated by signals of low energy supplies
ADP
AMP
FBPase-1 –
Inhibited by signals of low energy supplies
AMP

6. Hormonal regulation of glycolysis and gluconeogenesis
mediated by fructose 2,6 bisphosphate
F26BP activates PFK-1 (binds to allosteric
site and increases its affinity for its
substrate (F6-phosphate)
F26BP inactivates FBPase 1 (decreases
its affinity for its substrate (F1,6 BP))

7. **Regulation of NZ is by phosphorylation**
Where does F2,6 Bisphosphate come from?
Hormonal regulation of PFK-2 / FBPase-1
Formation of F26BP
Breakdown of F26BP
1 bifunctional NZ with 2 separate activities regulated by insulin and glucagon
**Regulation of NZ is by phosphorylation**
Glucagon stimulates phosphorylation of PFK-2/FBPase 2
-Activation of FBPase-2 activity (phosphatase)
-Reduction of F26BP
- Glycolysis Gluconeogenesis  
Insulin stimulates dephosphorylation of PFK-2/FBPase 2
-Activation of PFK2 activity (Kinase)
-Increase in F26BP
- Glycolysis Gluconeogenesis  

8. Enzymes of glycogen metabolism are regulated by allosteric
and hormonal mechanisms
Allosteric Regulation
Glycogen Synthase
Glycogen Phosphorylase
Activation G6P AMP
Inhibition ADP, Pi ATP, G6P, Glucose
Signals of high energy supply (G6P, ATP) stimulate glycogen synthesis and
inhibit glycogen breakdown.
Signals of low energy supply (ADP, AMP) stimulate glycogen breakdown
and inhibit glycogen synthesis

9. Inhibition of Glycogen
Hormonal Regulation of glycogen synthesis and breakdown – Covalent
Modification
Glycogen breakdown
Inhibition of Glycogen
breakdown
Glucagon
Activation of PKA (via cAMP)
Activation of phosphoprotein phosphatase -1
DeP of Glycogen phosphorylase
Phosphorylase kinase
P of phosphorylase kinase
P of Glycogen phosphorylase
And
P of Glycogen synthase
(A)
(IA)
(IA)
Stimulation of
glyc breakdown
Inhibition of glyc
synthesis
(A)
(IA)

10. ↓Glycolysis ↓Glycogen synthase Glycogen breakdown
Fed State - Insulin
Glucose enters hepatocytes through transporter
2) Synthesis of glycolytic enzymes
3) inactivation of GSK3 and activation of PP1
Activate glycogen synthase and inactivate glycogen
phosphorylase
Glycolysis
Glycogen synthase
↓Glycogen breakdown
Fasting State - glucagon
Activation of PKA through cAMP
1) Activates phosphorylase kinase  glycogen
phosphorylase
2) Inactivates glycogen synthase
3) Phosphorylates PFK-2/FBPase-2  in F2,6BP
4) Inactivates pyruvate kinase
↓Glycolysis ↓Glycogen synthase Glycogen breakdown

11. Cytoplasm Mitochondria Overview of Steps Glycolysis Glucose Pyruvate
Transition/Prep Phase
Mitochondria
Acetyl - CoA
NADH
FADH2
Citric
Acid
Cycle C1 C3
ATP
synthase C2 C4
ATP
Electron Transport

12. Citric Acid Cycle 2 C acetyl group combines with
4 C oxaloacetate to yield 6 C citrate 1-
1 Glucose
2 pyruvates
2- Carbons lost in pathway as C02
-These Cs are not from acetyl-coA
3- Oxaloacetate consumed in the 1st
step is regenerated in the last.
4- Energy produced is transferred as
energy-rich electrons to
NAD+ to yield NADH or to
FAD+ to yield FADH2
5- 2 rounds of Cycle yields:
4 CO2, 6 NADH, 2 FADH2, and
2 ATP
6- CAC is amphibolic – has a role in
oxidation of carbohydrates and provides
precursors for other pathways (ex – AA
metabolism)
Pyruvate Dehydrogenase
2 Acetyl-CoA
Citrate (C6)
Oxaloacetate (C4)
CoASH
Isocitrate (C6)
NADH
C02
Malate
NADH
a-ketoglutarate (C5)
Fumarate (C4)
CoASH
C02
FADH2
NADH
Succinate (C4)
CoASH
Succinyl co-A (C4)
Substrate level
phosphorylation
GTP / ATP

13. * * * * Regulation of Citric Acid Cycle
A- Substrate Availability (oxaloacetate;
acetyl-CoA)
Ca2
B- Product Inhibition
1- PD
C- Competitive Feedback Inhibition
NADH
Citrate
Regulated enzymes of CAC
4-CS
1- Pyruvate Dehydrogenase (irreversible)
Product/Feedback inhibition
Covalent Modification (Phosphorylation
= inactive)
2- ID *
2- Isocitrate Dehydrogenase
Product/
Feedback
Inhibition *
3- a-Ketoglutarate Dehydrogenase
3-KD *
4- Citrate synthase *
All function far from equilibrium (- ΔG)
NADH and Acetyl CoA compete
with NAD+ and CoA for binding sites –
competetive feedback inhibition

14. Electron Transport Chain

15. Matrix 2H+ 4H+ 4H+ Cyt. C Intermembrane space Complex I Complex III
Soluble
Coenzyme Q
FMN
FeS
Cyt’s
FeS
Cyt’s Cu
Complex II
Complex
III
Complex IV
FAD
FeS
Matrix
NADH
NAD+
FAD+
FADH2
½ O2 + 2H+
Succinate
H20
Fumarate
1- Electrons from NADH and FADH2 are passed through redox centers of 4 membrane bound complexes and 2 membrane soluble electron shuttles
2-During electron transfer, protons are translocated from the matrix into the
intermembrane space (Complex 1, Complex III, and Complex IV)
3- Final reduction is of molecular O2
4- Electrons from NADH yield 2.5 ATP; Electrons from FADH2 yield 1.5 ATP

16. Proton Motive Force - Chemiosmotic theory
Storage of energy as a proton/voltage gradient across a membrane
1- In ETC, the transport of H+ from low [H+] to high [H+] requires energy (ENDERGONIC)
2- Discharge of proton is EXERGONIC
Free energy of discharge of proton gradient is harnessed by ATP synthase
Outer membrane
Intermembrane
space
Low pH H+ H+ H+ Fo
++++++
++++++ l
lll lV V
Inner membrane H+
High pH
ATP F1
Oxidative phosphorylation

17. Electron transport and ATP synthesis are “coupled”
ATP synthesis requires discharge of proton gradient:
Proton gradient cannot be discharged without synthesis of ATP:
Proton gradient is established by electron transporting complexes.
No drug
uncoupler
added
Inhibitor added
DNP
DNP
DNP
ATP Synthesis
02 Consumption
Time
Time
Time
Uncoupler of ETC
Destroys proton gradient;
No stored energy for ATP;
DOES NOT stop electron flow;
Oxygen still being consumed
Electrons flowing through ETC;
O2 is being consumed;
ATP is being synthesized
Inhibitor of ETC
Stops flow of electrons;
No proton pumping;
No proton motive force;
No longer consuming O2;
No longer making ATP