ELECTRON TRANSPORT CHAIN

Lectures by Dr S.J. Brookes
If you have any questions relating to these lecture make an appointment for a Wednesday afternnon afternoons in the Division of Oral Biology (level 6). Alternatively you can email any problems to Dr. Brookes who will reply to your message as soon as possible (s.j.brookes@leeds.ac.uk).

Other pages related to this topic can be found at:

THCME FATTY ACID OXIDATION

UTDB FATTY ACID OXIDATION

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Introduction

The proton gradient across the inner mitochondrial membrane maintained by action of electron transport chain
Chain consists of 6 proteins associated with inner mitochondrial membrane:

  1. NADH dehydrogenase (complex I)
  2. Succinate coenzyme Q reductase (complex II)
  3. Coenzyme Q (CoQ) (also called ubiquinone)
  4. Cytochrome bc1 complex (complex III)
  5. Cytochrome c (Cyt c)
  6. Cytochrome oxidase (complex IV)

Electrons having a high reducing potential are removed from food molecules and are carried to the electron transport chain by electron carriers (NADH and FADH2). The details of how the carriers pick up these electrons will come later (see citric acid cycle and fatty acid oxidation) - for now just accept that they carry electrons and can act as reducing agents (reducing agents are molecules that can donate electrons).

STEP 1

NADH binds complex I & passes 2 electrons to a flavin momonucleotide (FMN) prosthetic group. The FMN is reduced to FMNH2. Each electron is transferred with a proton.
The electrons are then passed to iron-sulphur proteins (FeS) in complex I (this is non-heme iron). The electron is accepted by Fe3+ which is reduced to Fe2+:

(Remember that Fe3+ is reduced to Fe2+ by electrons)

STEP 2

Two electrons from the reduced FeS proteins are then passed to CoQ along with 2 protons. The CoQ is thus reduced to CoQH2 (ubiquinol) while the FeS proteins are oxidised back to Fe3+ state.


CoQ is small and lipid soluble so it is mobile in the mitochondrial membrane. It diffuses easily and shuttles the electrons to complex III (we will talk about complex II later).

STEP 3

Complex III contains cytochrome b, cytochrome c1 and FeS proteins. Like FeS proteins, cytochromes contain bound Fe atoms (this time the iron is heme). The iron atoms alternate between +3 and +2 oxidation states as they pass on the electrons.
CoQH2 passes 2 electrons to cyt b causing the Fe3+ to be reduced to Fe2+. The electrons are passed to the FeS protein and then to cyt c1.

STEP 4

Cyt c is another small mobile protein. It accepts electrons from complex III (Fe3+ is reduced to Fe2+) and shuttles them to the last electron transport protein in the chain (complex IV).

STEP 5

Complex IV contains cytochrome a and cytochrome a3 (both use Fe and Cu atoms to handle the electrons). Four cytochrome c molecules pass on 4 electrons to complex IV. These are eventually transferred with 4 H+ to O2 to form 2 water molecules.

This is a complex reaction mechanism and no attempt has been made in the above diagram to explain how the 4 electrons from 4 Cyt C are conveyed to the O2 (it doesn't balance with respect to electrons!)

What about complex II?

Complex II actually contains the enzyme succinate dehydrogenase which catalyses the reduction of succinate to fumarate (which as we will see later is a reaction of the so called citric acid cycle). FAD oxidises succinate to fumarate (FAD becoming reduced to FADH2 as it picks up 2 electrons and 2 protons). Succinate dehydrogenase is actually associated with complex II. FADH2 is oxidised back to FAD by passing the electrons on to FeS proteins in complex II. The electrons are then passed to CoQ and are passed on to complex III as described above in step 3.

Summary so far:

Electrons having a high reducing potential are removed from food molecules and are passed to electron carriers (NADH & FADH2). The electron carriers pass electrons onto the electron transport chain. The electrons are passed down the chain with each component being reduced as it accepts the electrons and re-oxidised as it passes them on. Eventually the electrons are used to reduce oxygen to water.

Essentially we have this reaction:

NADH + H+ + 1/2O2 NAD+ + H2O + ENERGY

The NADH is oxidised back to NAD+ by oxygen as the oxygen is reduced to water. This reaction releases a great deal of energy and would be useless to the cell if it was allowed to occur as written above. The reaction takes place via the electron transport chain which allows the energy to be released in a controlled way making it available to do useful work.

The energy is used to pump protons into the inter mitochondrial membrane space from where they flow down an electrochemical gradient. The energy released as the gradient dissapates is used to phosphorylate ADP to ATP (catalysed by ATP synthase).

How and where are protons pumped into inner membrane space?

NADH and FADH2 are powerful reducing agents (negative redox potentials) and readilly donate electrons to molecules having a more positive redox potential . Oxygen as a positive redox potential and readilly accepts electrons.

The redox difference between NADH and oxygen is +1.14 volts.

Oxidation of 1 NADH releases enough energy to make several ATP.


The 52.6kcal/mol is not released in one reaction but is released in small packets as each member of the electron transport chain reduces the next member. The bulk of the energy is released by 3 reactions involving coplexes I, III and IV. The energy is enough to transport 1 proton across the inner membrane at that point.

Proton pumping

Enough energy is released to pump 1 proton across the inner membrane as:

Electron transfers involoving CoQ and Cyt c do not release enough free energy to pump any protons. Proton pumping is not 100% efficient and some free energy is lost as heat during every step.

When FADH2 is oxidised by the electron transport chain the first proton pump (complex I) is bypassed since complex II has only enough reducing potential to pass electrons to CoQ.

Oxidation of NADH to NAD+ pumps 3 protons which charges the electrochemical gradient with enough potential to generate 3 ATP.
Oxidation of FADH2 to FAD+ pumps 2 protons which charges the electrochemical gradient with enough potential to generate 2 ATP.


Note: Allthough you will see these values quoted in many text books recent information suggests that 1NADH generates 2.5 ATP and 1FADH2 generates 1.5 ATP. The reason for this is that not all of the energy stored in the proton gradient is used to generate ATP. Some of the energy is used to power transport of ions in and out of the mitochondria.

Inhibitors of electron transport and ATP synthase


Inhibitors bind to the components of the electron transport chain and block electron transfer. All components before the block are stuck in a reduced state and all components after in an oxidised state. Clearly no electron transfer is possible and proton pumping stops. The proton gradient is quickly run down and ATP synthesis stops. Inhibitors may also block the proton channel (Fo) of ATP synthase. Such inhibitors are clearly toxic as ATP is essential to all living processes.

Inhibitor

Action

Cyanide, carbon monoxide

Blocks complex IV

rotenone, amytal

Blocks complex I

antimycin

Blocks complex III

oligomycin

Blocks the proton channel (Fo) in ATP synthase

(These inhibitors are useful tools used to study the electron transport chain).

Uncoupling agents

These make the inner membrane permeable to protons. The proton gradient leaks away bypassing the ATP synthase (electron tramsport has been uncoupled from ATP synthesis). The energy released is wasted as heat instead of being stored as ATP.

e.g. 2,4-Dinitrophenol (DNP) is lipid soluble so it can diffuse across the inner membrane. DNP is protonated while it is in the inner mitochondrial space (since the pH is relatively low due to the high proton conc.) It diffuses across the inner membrane into the matrix where the proton is released (since the pH is higher). The proton has crossed the inner membrane but has bypassed the ATP synthase and no ATP has been generated.

Uncoupling agents also occur naturally. New born and hibernating animals contain brown fat. Brown fat mitochondria contain the protein thermogenin which provides a channel through the inner mitochondrial membrane. The heat energy released as the protons rush down their concentration gradient through this channel keeps the animal warm.

Summary

Reduced electron carriers NADH & FADH2 reduce oxygen to water via the electron transport chain. The energy released is used to set up a proton gradient across the inner mitochondrial memebrane. The protons flow down this concentration gradient back across the inner mitochondrial membrane through the ATPase (Fo channel). The energy released is used to generate ATP.

WHERE DOES THE NADH AND FADH2 COME FROM?

The citric acid cycle and fatty acid oxidation supply NADH and FADH2.