oxidative phosphorelation

1. An Introduction to Metabolism and oxidative
phosphorelation
Assist prof. Dr.
Abdulhussien Aljebory
Babylon University / College of Pharmacy
Biochemistry 2016

2. General Considerations
 How do we define oxidative phosphorylation?
formation of ATP using the energy released by the transfer of electrons
from NADH and FADH2 through a series of electron carriers
 What couples the formation of ATP to the transfer of electrons?
a proton gradient
 Where in the cell does oxidative phosphorylation take place?
inner mitocondrial membrane
 What do we know about the mitocondrial membranes?
outer membrane – reasonably permeable
contains porins – VDAC (Voltage Dependent Anion Channel)
inner membrane – relatively impermeable

3. An Introduction to Metabolism
Catabolic Pathways
Release energy by breaking down complex molecules into
simpler ones.
Cellular respiration provides energy for cellular work.
C6H12O6 + 6O2  6CO2 + 6H2O + energy
Energy released drives anabolic reactions.
Anabolic Pathways
Consume energy by building molecules
Photosynthesis uses energy
6CO2 + 6H2O energy C6H12O6 + 6O2

4. Organisms Transform Energy
Solar
Energy
(EK)
Plants (glucose)
Stored in
chemical bonds
(EP)
Animals
Break down
Sugars;
Some used (EK),
some stored in
chemical bonds
(EP)

5. Type of metabolic pathways:
 Catabolic pathways release energy by breaking down complex molecules into
simpler compounds
 Anabolic pathways consume energy to build complex molecules from simpler
ones
 Bioenergetics is the study of how organisms manage their energy resources
 Metabolism:The sum of all the chemical processes occurring in an organism at
one time
 Concerned with the management of material and energy resources within the cell
 Catabolic pathways
 Anabolic pathways

6. Energy
 Kinetic energy is energy associated with motion
 Heat (thermal energy) is kinetic energy associated with random movement of atoms or molecules
 Potential energy is energy that matter possesses because of its location or structure
 Chemical energy is potential energy available for release in a chemical reaction
 Energy can be converted from one form to another
Laws of Thermodynamics
First Law—Energy can be transferred, but never created or destroyed
Second Law—Every energy transfer results in increased entropy
(randomness in the universe)
Some of the energy is converted to heat
Reactions occur spontaneously

7. The First Law of Thermodynamics
According to the first law of thermodynamics, the
energy of the universe is constant
 Energy can be transferred and transformed
 Energy cannot be created or destroyed
The first law is also called the principle of
conservation of energy
 The Second Law of Thermodynamics:
 During every energy transfer or transformation, some energy is unusable, often
lost as heat
 According to the second law of thermodynamics, every energy transfer or
transformation increases the entropy (disorder) of the universe
 10% Rule
 Entropy is a measure of disorder, or randomness

8. Biological Order and Disorder
 Cells create ordered structures from less ordered materials
 Organisms also replace ordered forms of matter and energy with less ordered
forms
 Entropy (disorder) may decrease in an organism, but the universe’s total entropy
increases
 A living system’s free energy is energy that can do work when temperature and
pressure are uniform, as in a living cell

9. Free Energy
 Organisms live at the expense of free energy (portion of a system’s energy available for work)
acquired from the surroundings
 Free energy is needed for spontaneous changes to occur.
 Gibbs-Helmholtz Equation
 G = H - TS
 Can be used to determine if a reaction is spontaneous
 Spontaneous reactions occur in systems moving from instability to stability
Total
energy
enthalpy
Free
energy
Temp
(K)
entropy
High
energy
Low
energy

10. Gibbs-Helmholtz Equation
 In chemical reactions, reactions absorb energy to break bonds
 Energy is then released when bonds form between rearranged atoms of the
product
 Indicates amount of energy available for work
 Indicates whether a reaction will occur spontaneously (low G)
 G decreases as reaction approaches equilibrium
 G increases as reaction moves away equilibrium
 G = 0 when a reaction is in equilibrium
G =  H - T  S
Measure
of heat
in the
reaction

11. (a) First law of thermodynamics (b) Second law of thermodynamics
Chemical
energy
Heat

12. Chemical Reactions
Exergonic Endergonic
Chemical products have lower G
than reactants
Products store more G than
reactants
Reaction releases energy Reaction requires energy input
(absorbs)
G = negative value G = positive value
Spontaneous Non spontaneous
In cellular metabolism, exergonic reactions drive
endergonic reactions

13. Reactants
Energy
Products
Progress of the reaction
Amount of
energy
released
(G < 0)
Freeenergy
Exergonic reaction: energy released

14. Reactants
Energy
Products
Progress of the reaction
Amount of
energy
required
(G > 0)
Freeenergy
Endergonic reaction: energy required

15. ATP couples exergonic reactions to
endergonic reactions
 A cell does three main kinds of work:
Mechanical
Transport
Chemical
 To do work, cells manage energy resources by energy coupling, the use
of an exergonic process to drive an endergonic one

16. ATP Powers Cellular Work
Unstable
Bonds—can release
energy when
broken
Energy transferred to
another molecule (phos-
phorylated intermediate)
with the phosphate
Less stable More stable

17.  Energy is released from ATP through the loss of phosphate groups
 Catabolic reaction resulting from hydrolysis producing ADP + Pi
(inorganic Phosphate) + energy (G = -7.3Kcal/mol in the lab,
-13Kcal/mol in the cell)
 How ATP works:
 Hydrolysis of ATP produces inorganic phosphate that is attached to
a molecule involved in an endergonic process
 Phosphorylation is the process of ATP transferring phosphate to a
molecule
 Results in a phosphorylated intermediate that can complete the
intended reaction

18. Enzymes
 Catalyst—chemical agent that speeds up a chemical reaction without
being consumed by the reaction
Hydrolysis of sucrose by the enzyme sucrase is an example of an
enzyme-catalyzed reaction
Sucrose
C12H22O11
Glucose
C6H12O6
Fructose
C6H12O6

19. The Activation Energy Barrier
 Every chemical reaction between
molecules involves bond breaking
and bond forming
 The initial energy needed to start a
chemical reaction is called the free
energy of activation, or activation
energy (EA)
 Activation energy is often supplied
in the form of heat from the
surroundings
Transition state
C D
A B
EA
Products
C D
A B
G < O
Progress of the reaction
Reactants
C D
A B
Freeenergy

20. Enzymes
 Catalytic proteins that speed up metabolic reactions by
lowering energy barriers
1. Reactants must absorb
energy to reach transition
state (unstable)
2. reaction occurs and energy
is released as new bonds
form to make products
3. G for overall reaction is
difference between G of
products and G of reactants

21. Substrate Specificity of Enzymes
 Substrate—reactant that an enzyme acts
 Substrate binds to the active site on the enzyme
 Induced fit of a substrate brings chemical groups of the active site into positions
that enhance their ability to catalyze the reaction.
How do Enzymes Work?
Active site holds 2 or more reactants in the proper position to react
Induced fit may distort chemical bonds so less thermal energy is needed
to break them
Active site may provide micro-environment that aids a reaction (localized
pH)
Side chains of amino acids in active site may participate in reaction

22. Control of Metabolism
 Allosteric Regulation: enzyme function may be stimulated or inhibited by attachment of
molecules to an allosteric site
 Feedback Inhibition: end product of metabolic pathway may serve as allosteric inhibitor
 Cooperativity: single substrate molecule primes multiple active sites increasing activity
 Equilibrium and Metabolism
 Reactions in a closed system eventually reach equilibrium and then do no
work
 Cells are not in equilibrium; they are open systems experiencing a constant
flow of materials
 Dynamic Equilibrium
 A catabolic pathway in a cell releases free energy in a series of reactions

23. ATP powers cellular work by coupling exergonic reactions
to endergonic reactions
A cell does three main kinds of work:
 Mechanical, Transport and Chemical
To do work, cells manage energy resources by
energy coupling, the use of an exergonic process
to drive an endergonic one
ATP (adenosine triphosphate) is the cell’s energy
shuttle
ATP provides energy for cellular functions

24. How do living organisms create macromolecules, organelles,
cells, tissues, and complex higher-order structures?
 The laws of thermodynamics do not apply to living organisms.
 Living organisms create order by recycling and reusing energy from the sun.
 Living organisms create order locally, but the energy transformations generate
waste heat that increases the entropy of the universe.

25. If this is an enzyme-catalyzed reaction, how can the rate of this
reaction be increased beyond the maximum velocity in this
figure?
 Increase the substrate concentration.
 Increase the amount of enzyme.
 Raise the temperature to be more
optimal.
 B is the best choice, but A and C are
also possible.
 There is no way to increase the rate of
the reaction any further.

26.  Viox is a nonsteroidal anti-inflammatory drugs (NSAIDs) are potent
inhibitors of the cyclooxygenase-2 (COX-2) enzyme. High substrate
concentrations reduce the efficiency of inhibition by these drugs. These
drugs are:
 competitive inhibitors.
 noncompetitive inhibitors.
 allosteric regulators.
 prosthetic groups.
 feedback inhibitors.

27. How does the flow of energy through life differ from the flow of matter
through life?
 Matter can be recycled, while some energy is always converted to
unusable forms like heat.
 Matter is brought into life from outside, while energy is generated from
within life.
 Life is able to convert energy into matter, through photosynthesis.
 Matter is conserved, while life causes energy to be lost over time.
 Life uses the flow of matter to keep its energy state unbalanced.

28. Redox Potentials and Free Energy Changes
 What is the relationship between change in redox potential and change in free energy?
G01 = -nF E1
0
n = number of electrons transferred
F =faraday (constant, 23.06 kcal/mole/volt)
Can calculate free energy change from reduction potentials of the
reactants
 By knowing the electron transfer potential of NADH relative to O2 one can calculate the
amount of free energy released when O2 is reduced by NADH.
 One can also quantify the energy associated with a proton gradient.
 G = RTln(c2/c1) + ZF V
c2 = concentration on one side of membrane
c1 = concenetration on other side of membrane
Z = electrical charge of transported material
F = Faraday constant (23.06 kcal/mole/volt)

29. Electron Transport
What are the complexes making up the
respiratory chain?
three proton pumps
one link to citric acid cycle

30. Electron Transport - conyinue
Initial step is transfer of electrons to FMN a prosthetic group of the
enzyme
Electrons are then transferred to iron-sulfur clusters another
prosthetic group

31. Electron Transport - continue
 Electrons from clusters transferred to coenzyme Q
 as a result of electron transfer four protons are pumped out of mitocondrial matrix
 Reaction summarized:
NADH + Q + 5H+
matrix  NAD+ + QH2 + 4H+cytosol
 Coenzyme Q also serves as entry point for electrons from FADH2 from oxidation of succinate
 succinate-Q reductase complex
 inner mitocondrial membrane
 FADH2 transfers electrons to iron-sulfur clusters then to Q
 no protons are pumped
 Q-cytochrome c oxidoreductase catalyzes the transfer of electrons from Q to cytochrome c
 What is a cytochrome?
 electron transferring protein with heme prosthetic group
 transfers only electrons
 iron in heme goes between Fe+2 and Fe+3

32. Electron Transport
Q-cytochrome c oxidoreductase contains
3 hemes and a iron-sulfur cluster
 What is the function of cytochrome c oxidase?
reduction of oxygen to water
 What are the major prosthetic groups of this
complex?
CuA/CuA
heme a
heme a3-Cub

33. Electron Transport- continue
 Toxic derivatives of molecular oxygen may be formed by partial
reduction
O2 O2
_
O2
_
2
 How does the cell protect itself against these reactive oxygen species?
makes use of superoxide dismutase and catalase
2O2
_
+ 2H+  O 2 + H2O2
2H2O2  O2 + 2H2O
Superoxide anion peroxide

34. ATP Synthesis
What is the chemiosmotic hypothesis?
ATP synthesis and electron transport are coupled by proton gradient across
mitocondrial membrane
What is ATP synthase and
what do we know about
its structure?
consists of F1 and F0
F1 has 5 types of polypeptide chains
3,3,,,
F0 contains proton channel
10-14 c subunits
a,b2 subunits

35. ATP Synthesis
 How does proton flow through F0 drive the rotation of the 
subunit?
 each c subunits consists of 2  helices with one helix containing an
aspartic acid residue
 a subunit contains two proton half channels
 Proton enters half-channel, neutralizes charge on aspartate
 c can rotate clockwise
 proton can move into matrix
 Since c ring is linked to  and  subunits, as c turns these subunits rotate
 rotation protmotes synthesis of ATP via binding-change mechanism
 each 3600 rotation of  subunit leads to synthesis of 3 ATP’s
10 protons generate 3 ATP’s
each ATP requires transport of about 3 protons

36. Regulation of Respiration
Oxidative phosphorylation
can be inhibited by many
substances