Download Cytochromes

BIOLOGICAL OXIDATION
Lecture Objectives;
At the end of lecture Student will be able to:
►
Explain the fundamentals of Cellular Respiration.
►
Describe the biochemical mechanisms of generation and utilization of
energy.
►
Describe the effects of various inhibitors on biological oxidation.
► Study
Bioenergetics
(Biochemical Thermodynamics)
of the energy changes accompanying biochemical
reactions.
► Study of generation, storage and utilization of energy in living
system.
PRODUCTION OF ENERGY IN A LIVING
S Y S TE M
• In living system energy is generated by precise chemical
methods which:
Is Precisely controlled and regulated according to requirements
Can be stored for use at a proper time
Do not require drastic conditions of Temperature, Pressure and pH.
► Medical




Biomedical importance
problems due to energy imbalance:
Starvation
Obesity
Marasmus
Diseases of Thyroid
Sources of energy in living system
► The overall process of energy generation by breakdown of
food is called Cellular Respiration
► It is the process by which the chemical energy of "food“
molecules is released and captured in the form of ATP.
► Carbohydrates, fats, and proteins can all be used as fuels in
cellular respiration.
► Process involves oxidation of energy-rich organic compounds
coupled with reduction of other compounds.
► Direct At Substrate Level.
► Indirect Through Biological Oxidation.
Storage of energy in living system
► Free or useful energy is released during catabolic reactions.
► This energy is stored as high energy phosphate bonds of
various molecules.
► Such molecules are called high-energy compounds.
Adenosine Triphosphate (ATP) is the most commonly
used Energy Storing Molecule
► Adenosine
+ ribose + P ___P___P (High Energy Bonds)
Hydrolysis of terminal “P” generates 7300 calories
ATP is converted to ADP.
Reduced Coenzymes (NADH & FADH) are oxidized to
generate Energy
► NADH & FADH are generated during Glycolysis, Oxidation of Fatty Acids,
Catabolism of Amino Acids and TCA cycle.
► Oxidized in mitochondria to generate ATP.
Two types of Cellular Respiration
► Aerobic Respiration.
► Anaerobic Respiration.
Aerobic Respiration
► Final Oxidizing agent (Electron Acceptor) is Oxygen
► The most Important Metabolic Source of Energy in
Living cells
► Contributes approximately 90% of total energy generated
in cell.
► Net result is CO2, H2O and Energy (ATP & Heat)
Anaerobic Respiration
► Final Oxidizing agent (Electron Acceptor) is Other
than Oxygen.
► Important Source of Energy in:
 Cells Not Having Mitochondria (RBC).
 Cells Deprived of Oxygen (Exerting Muscles &
Ischemic Tissues)
Metabolic Processes Involved
In Cell Respiration
► Glycolysis; occurs in cytosol.
► Oxidation of Fatty acids and
Krebs Citric acid Cycle occur
in Matrix of Mitochondria.
► Biological Oxidation occurs in Inner Membrane of
Mitochondria.
 Carbon skeleton of Glucose, Fatty acids and Amino acids are
converted to CO2 along with generation of NADH2 and FADH
molecules.
► Final stage of cellular respiration
► It is the oxidation of NADH2 & FADH to generate ATP
► Generates 90% of total energy generated by Cellular
Respiration
Glucose Metabolism
Glycolysis
HIGH ENERGY COMPOUNDS
Glucose

Glyceraldehyde 3-P

1,3 DPG

3, Phosphoglycerate

PEP

Pyruvic acid.
NAD →NADH2
ADP→ATP
Fatty Acid Metabolism
LIPOLYSIS
Fatty acid

Acyl~CoA

Unsaturated Acyl~CoA

 -Keto Acyl~CoA

Acetyl~CoA
HIGH ENERGY COMPOUNDS
FAD→FADH2
NAD→NADH2
Amino Acid Metabolism
► Amino Acids are catabolized, after removal of amino group,
to generate Intermediates of TCA Cycle
 Glutamic acid-Ketoglutaric Acid
 Aspartic AcidOxaloacetic Acid
 PhenylalanineAcetyl CoA
► These Intermediates generate NADH & FADH through Citric
Acid Cycle.
Mechanism of Biologic Oxidation
► Includes
–
–
two integrated mechanisms
Respiratory Chain: extracts energy from electrons of Hydrogen.
Oxidative Phosphorylation: utilizes extracted energy for the
formation of high-energy bond.
Respiratory Chain;
Electron Transport Chain (ETC)
► Electrons carried by reduced NAD and FAD are transported
finally to Oxygen (for oxidative phosphorylation) through
a gradual downward movement, from high energy level to
low energy level.
► A series of Proteins & enzymes Complexes, cytochromes,
present in inner membrane of mitochondria, act as
temporary electron carriers during downward movement of
electron.
► Movement from high to low level librates energy that
is utilized to generate ATP.
Cytochromes
► Membrane bound hemoproteins.
► Act as electron carrriers.
► Contain heme (iron porphyrin) as Prosthetic Group.
► In
Types of Cytochromes
Human Mitochondria Cytochromes Related to
Respiration are Distinguished As:




Cytochrome b
Cytochrome c
Cytochrome c1
Cytochrome a and Cytochrome a3
Two Categories of electron
carriers (Components Of ETC)
1. Large, immobile Protein complexes.
Complex I
Complex II
Complex III
Complex IV
2. Mobile electron carriers.
Coenzyme Q
cytochrome C
Oxygen
Large, Immobile Protein Complexes of
Respiratory Chain
► E m b ed d ed




in Inner Mitochondrial Membrane:
Complex I; NADH dehydrogenase.
Complex II; Succinate-ubiquinone oxidoreductase.
Complex III; the bc1 complex or ubiquinol-cytochrome c reductase.
Complex IV; Cytochrome Oxidase.
Mobile electron carriers of
Respiratory Chain
►Shuttles electrons between Complexes.
►Serve as links between ETC complexes:
 Ubiquinone (Coenzyme Q).
 Cytochrome c.
Cytochromes
Found in two different environments
1. The mobile cytochrome electron carriers (Such as
Cytochrome c).
2. Subunits of bigger enzymatic complexes such as Cyt. b in
Complex III. and Cyt. a, a3 in Complex IV.
►
Step #1:
Steps of Electron Transport
 Transfer of Electron to complex I and complex II.
► Step
#2 :
► Step
#3 :
► Step
#4 :
 Transfer of Electron to Coenzyme Q.
 Coenzyme Q shuttles electrons and pass them to complex III.
 Transfer of Electron to Cytochrome c.
► Step
#5
► Step
#6 :
 Cytochrome c shuttles electrons and pass them to complex IV.
 Transfer of Electron to Oxygen.
Net result of ETC
► Recovery of Free NAD & FAD:
 Oxidized (Free) NAD & FAD are recovered to Continue
dehydrogenation reactions of catabolism.
► Formation of water:
 NADH2 + ½ O2
NAD+ + H2O
 FADH2 + ½ O2
FAD+ + H2O
► Energy which is captured as ATP by process of Proton Pump.
Flow Of Electrons through Complexes I, III and IV release energy
which is used to pump protons across the IMM and from a
"Proton Gradient”
High Proton Gradient
In Inter-membrane space
[H+]
NADH
FMNH2
[H+]
QH2
Cyt. b
Cyt. c
Cyt. a-a3
[H+]
O-
Mitchell’s Hypothesis
The Chemiosmotic Hypothesis for oxidative
phophorylationElectron transport through the ETC
generates a proton concentration gradient (Proton
motive force; PMF).
► PMF serves as the energy reservoir for driving ATP
formation.
► This energy is utilized for the synthesis of ATP, when
Protons flow back into the matrix via ATP synthase (Complex
V).
► Chemiosmotic Potential or Proton-Motive Force (PMF) The
electrochemical potential difference between the two sides
of the IMM, that engage in active transport of Protons is
called Proton-Motive Force (PMF).
► Proton motive force is the energy of the proton
concentration gradient for OXIDATIVE PHOSPHORYLATION.
► Proton flow back to matrix “Proton Channel” & Release
energy for synthesis of ATP
► ATP Synthase (Complex V) has 2 components: F1 and F0
Mitochondrial Chemiosmosis; H+ flow
forms a circuit (similar to an electrical
circuit)
Estimation Of ATP Synthesis
► According to most textbooks:
 3 ATPs for NADH and 2 ATPs for FADH.
► According to recent research:
 10 protons are pumped by NADH.
 6 protons are pumped by FADH.
 4 Protons are needed by ATP synthase to make one
ATP molecule.
 This means that each NADH can make 2.5 ATPs
(10/4) and each FADH can make 1.5 ATPs (6/4).
FORMATION OF SUPEROXIDE
Complex I & III are the main sites of premature electron
leakage to oxygen, producing free radical superoxide, which
contributes to pathology of different diseases and aging
UNCOUPLERS
In some special cases, Uncouplers stimulate the oxidation of
substrates in the absence of ADP so that large amounts of O2 are
consumed but no ATP is produced.
► Do NOT affect electron transport
Agents that Interfere With ATP
Synthesis
► GENETIC DEFECTS
► Respiratory Chain Inhibitors:
 CN, CO, Antimycin A etc.
► ATP Synthase Inhibitors:
 Oligomycin.
► Uncouplers:
 DNP (synthetic) and Thermogenin(natural).
 ATP-ADP Exchange Inhibitors: Atractyloside.