2. Electron transport chain (ETC) and its mechanism.
Oxidative phosphorylation & its mechanism
Substrate level phosphorylation
Inhibitors ETC and oxidative phosphorylation
/Uncouplers
3. Oxidation is the loss of electrons and reduction is the gain of
electrons.
oxidation-reduction is applicable to biological systems also.
The transfer of electrons from the reduced coenzymes through
the respiratory chain to oxygen is known as biological
oxidation.
The enzymes involved in biological oxidation are:
1. Oxidases
2. Dehydrogenases
3. Hydroperoxidases
4. Oxygenases.
4. Energy-rich molecules, such as glucose, are metabolized by a
series of oxidation reactions and yield CO2 and water.
The metabolic intermediates of these reactions donate
electrons to specific coenzymes, nicotinamide adenine
dinucleotide (NAD+) and flavin adenine dinucleotide (FAD), to
form the energy-rich reduced forms, NADH and FADH2.
These reduced coenzymes donate a pair of electrons to a
specialized set of electron carriers, collectively called the
electron transport chain
The ETC is located in the inner mitochondrial membrane.
The inner mitochondrial membrane have 5 enzyme complexes-
complex I, II, II , IV and V.
The complexes I- IV are carriers of electrons while complex V
is responsible for ATP synthesis.
5. There are five distinct carriers that participate in the ETC.
(1)Nicotinamide nucleotides:
NAD+ is reduced to NADH + H+ by dehydrogenases with the removal
of 2 hydrogen atoms from the substrates (glyceraldehyde-
3phosphate, pyruvate,isocitrate,α-ketoglutarate and malate).
(2) Flavoproteins:
NADH dehydrogenase reductase is a flavoprotein with FMN as the
prosthetic group.
The coenzyme FMN accepts 2 electrons and a proton to form
FMNH2.
Succinate dehydrogenase is also a flavoprotein with FAD as the
coenzyme. This accept 2 hydrogen atoms from succinate
(3) Iron- sulfur proteins:
The iron-sulfur (FeS) proteins exist in the oxidized (Fe3+) or
reduced (Fe2+) state and they participates in the transfer of
electron
6. (4) Coenzyme Q (ubiquinone):
It can accept electrons from FMNH2 produced in the ETC by
NADH dehydrogenase or FADH2 produced outside ETC
(5) Cytochromes:
cytochromes are conjugated proteins containing heme group
The electrons are transported from coenzyme Q to cytochromes
(in the order) b, c1, c, a and a3.
8. The transport of electrons through the ETC is linked with the
release of free energy.
The process of synthesizing ATP from ADP and Pi coupled with
the ETC is known as oxidative phosphorylation.
The complex V of the inner mitochondrial membrane is the
site of oxidative phosphorylation.
There are three reactions in the ETC that are exergonic to
result in the synthesis of 3 ATP molecules.
1. Oxidation of FMNH2 by coenzyme Q.
2. Oxidation of cytochrome b by cytochrome c1 .
3. Cytochrome oxidase reaction.
9. MECHANISM OF OXIDATIVE PHOSPHORYLATION.
Chemical coupling hypothesis :
According to chemical coupling hypothesis, during the course of
electron transfer in respiratory chain, a series of phosphorylated
high-energy intermediates are first produced which are utilized
for the synthesis of ATP.
Chemiosmotic hypothesis :
Proton gradient : The inner mitochondrial membrane is
impermeable to protons (H+) and hydroxyl ions (OH-).
The transport of electrons through ETC is coupled with the
translocation of protons (H+) across the inner mitochondrial
membrane (coupling membrane) from the matrix to the
intermembrane space.
The pumping of protons results in a proton gradient which results
in the synthesis of ATP from ADP and Pi.
ATP synthase, present in the complex V, utilizes the proton
gradient for the synthesis of ATP.
10. Rotary model for ATP generation:
The changes in the mitochondrial membrane proteins leads
to the synthesis of ATP.
Structure of mitochondrial ATP
synthase complex
Inner mitochondrial
membrane
11. ATP may be directly synthesized during substrate oxidation
in the metabolism.
The high-energy compounds such as phosphoenolpyruvate
and 1,3-bisphosphoglycerate (intermediates of glycolysis)
and succinyl CoA (of citric acid cycle) can transfer high-
energy phosphate to ultimately produce ATP.
12. (1)NADH and coenzyme Q :
Fish poison rotenone, barbituate drug Amytal and antibiotic
piercidin A inhibit this site.
(2)Between cytochrome b and c1 :
Antimycin A -an antibiotic, British antilewisite (BAL)-an
antidote.
(3)Inhibitors of cytochrome oxidase:
Carbon monoxide, cyanide, hydrogen sulphide and azide
effectively inhibit cytochrome oxidase.
Carbon monoxide reacts with reduced form of the cytochrome
while cyanide and azide react with oxidized form.
Cyanide is a potent inhibitor of ETC. It binds to Fe3+ of
cytochrome oxidase blocking mitochondrial respiration leading
to cell death.
13.
14. Uncouplers are compounds that can uncouple (or delink) the
electron transport from oxidative phosphorylation
They increase the permeability of inner mitochondrial
membrane to protons (H+), so that ATP synthesis does not
occur.
The uncouplers allow oxidation of substrates without ATP
formation.
Eg: 2,4-dinitrophenol (DNP), dinitrocresol, pentachlorophenol,
trifluorocarbonylcyanide phenylhydrazone( FCCP).
Physiological substances like thermogenin, thyroxine and long
chain free fatty acids
The antibiotic Oligomycin prevents the mitochondrial oxidation
as well as phosphorylation.
Plant toxin Atractyloside is an uncoupler