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Biochemistry Home Page
Teaching Graduate Medical Course - Biochemistry Page Primers, Readers, and Reading Lists page Oxidative phosphorylation Learning Resource| Author(s)... |
| Dr Peter Nixon |
| Revised... |
| 8th March 2000 |
| Downloads... |
| as PDF file...
as Word6 file... |
| Keywords... |
Students with no background in Biochemistry will want this introduction and textbook reading list on the topic of oxidative phosphorylation. Students with a background in Biochemistry will probably want to review the topic before attempting the questions. Students with a very strong background in Biochemistry might like to go straight to the questions and then decide how much you need to review.
This SDL will help provide an understanding of the molecular basis of the obligatory requirement of almost all cells for a constant supply of oxygen, and hence to for an uninterrupted blood supply. Necessarily, it introduces the concepts of energy, particularly chemical energy; energy "currency" within cells as energy-rich chemicals; oxidation and reduction; subcellular structures specialised to separate components of an electrochemical gradient; electron-transfer molecules; and electron transfer to oxygen.
Initially, you should look for answers to such questions as the following:
In review, you will want to apply your knowledge of oxidative phosphorylation to practical questions; the concluding questions allow that application. You will also be able to apply your knowledge of oxidative phosphorylation in the practical laboratory in week 4 of the GMC.
Blood must circulate for many reasons: two major ones are to transport oxygen from the lungs to other tissues and carbon dioxide in the reverse direction. Any interruption of this constant process is lethal to the affected cells. Why ?
Food is utilised, in part, to provide energy in order to maintain a highly ordered metabolic state and to carry out useful work. Energy is harnessed principally by the process of oxidative phosphorylation, an aerobic, or oxygen-requiring, process. In fact, not all energy-yielding reactions require oxygen, but those linked to oxygen consumption are so much more efficient than those independent of oxygen that, in practice, more than 85% of our energy requirements are met by the process of oxidative phosphorylation. The remaining energy is derived from incomplete oxidation of foods by reactions that do not involve oxygen. The reading list focusses on the process of oxidative phosphorylation; this introduction serves to define necessary underlying concepts.
Energy is consumed when work is done; that is, energy is the potential to do work. Work and energy share the same dimensions and units; in the case of heat and chemical energy the units are the joule in S.I. or the calorie in older literature. At the molecular level, the breaking and making of chemical bonds are processes that require an energy input. The energy required to form a chemical bond is a characteristic of each particular class of bond, and differs between classes. One chemical bond requiring a rather high "free energy" input for its formation, or releasing the same high "free energy" on its cleavage, is the terminal phosphate ester bond of a triphosphate, such as adenosine triphosphate (known by the abbreviation ATP), or guanosine triphosphate (GTP). Pages 192-193 of Dow, Lindsay and Morrison "Biochemistry", 1996, describe and illustrate the basis of the energy status of the bonds linking phosphate groups in ATP.
All living organisms couple the oxidation of foods to the generation of ATP by phosphorylation of adenosine diphosphate (ADP), then utilise the energy contained within the terminal phosphate ester bond to drive other chemical reactions which are themselves energy-requiring, at the same time breaking the terminal phosphate bond to produce ADP again. The energy released by the hydrolysis of ATP to produce ADP is 30.5 kJ/mol. Thus ATP is continually synthesised and broken down, as required, and resynthesised, sustained by a small total body amount of the sum of ADP and ATP. In fact the turnover of ATP within a sedentary 70 kg human amounts to 40 kg per 24h, and can be as high as 0.5 kg per minute during strenuous exercise!
The work done by ATP is varied and includes activation steps in the synthesis of other compounds needed by cells, such as proteins and nucleic acids (both macromolecules requiring much energy for their synthesis); the driving of transmembrane pumps, which maintain the intracellular and extracellular chemical environments upon which human life depends; reactions in the recognition of hormonal signals; and the generation of motion, such as muscle contraction or the movement of molecules between compartments within cells. The work done by a single human heartbeat is approximately 0.5 J, requiring the synthesis and breakdown of ATP. The daily requirement of energy to drive the contractions of the human heart (in excess of 100,000 per day) then amounts to more than 50 kJ, requiring the hydrolysis of more than 0.8 kg of ATP or 2% of daily energy expenditure of a sedentary adult.
Fossil fuels can be burnt in oxygen only at high temperature, but the reaction releases much energy and other products. If the fuel is a pure hydrocarbon, the other products are carbon dioxide and water, and the release of energy can be explosive. If the fuel combustion is confined, for example by an engine, the released energy can be harnessed to do work, for example to propel a vehicle, but with rather low efficiency: perhaps only 30% of the available energy can be utilised, the remainder being lost as heat. The marvel of energy extraction from foods by living organisms is that it can be achieved at a low temperature (approximately 37o in the case of man) under precise control, modest efficiency and without even a hint of an explosion.
This is achieved by controlled sequences of chemical reactions which progressively oxidise foods in small steps, simultaneously creating a reduction potential (more exactly, usually reducing the molecule nicotinamide adenine nucleotide, the reduced form of which is known by the abbreviation NADH. [The structure of the oxidised form NAD+, is on pp 187-8 of Dow et al]. One byproduct of the oxidation of foods is carbon dioxide, which is excreted by the lungs. In later weeks we will return to these processes and how they are controlled. In mitochondrion NADH is split first into NAD+ and the hydride ion, H- ; the hydride ion is separated into a proton (H+ ) and 2 electrons (e- ). The electrons are transferred in small steps to successive electron carriers (the oxidation part of oxidative phosphorylation) and ultimately to oxygen with the reduction of oxygen to water, a process which simultaneously drives the proton across the inner membrane of the mitochondrion to generate a transmembrane proton gradient, and which is coupled to the phosphorylation of adenosine diphosphate (ADP) to synthesise adenosine triphosphate (ATP - the phosphorylation part). Since major features of this process are the transfer of electrons to oxygen and the coupled phosphorylation of ADP to ATP, it is known as "oxidative phosphorylation".
The reading list directs you to consider this mitochondrial process, assuming the reactions responsible for the reduction of NAD to NADH, which you will meet later. The rate of ATP synthesis varies according to need; by studying oxidative phosphorylation we understand not only how aerobic processes yield useful energy from nutrients but also why people lose weight when they diet and why they breath more heavily when exercising.
Read the outline of this topic in Dow, Lindsay and Morrison (see below) then progress to one of the other listed textbooks, depending on your previous level of exposure to biochemistry. Lehninger et al. is recommended.
A brief overview account which is clear and well-illustrated and is recommended both for review by those who have already studied Biochemistry, and as a definitive introduction for those who have not. The material on pp. 194-198 is somewhat peripheral this week but will be taken up in later weeks.
Thermodynamics of Biological Systems, Chapter 3, pp56-80,
Metabolism and its regulation, Chapter 18, pp565-608
Electron Transport and Oxidative Phosphorylation, pp 673-708.
An excellent description, lavishly illustrated and very readable; perhaps the best account and up-to-date. Chapter 3 establishes the ideas of energy while chapter 18 establishes the ideas of metabolism and links those to nutrition. Chapter 21 gives the account of oxidative phosphorylation. This book is complemented by the book and CD-ROM Interactive Biochemistry by Grisham; the CD-ROM contains an interactive video depicting ATP synthase in motion. This book and CD are in the Biological Sciences Library. and are highly recommended.
A clear and direct account is provided by these authors, perhaps partly because their principal research interest is oxidative phosphorylation. For this topic, this text is highly recommended for revision by those who have previous Biochemistry and for those without that background who cope well reading beyond the account of Dow et al. Chapter 13, Principles of Bioenergetics (pages 364 - 397) provides the necessary introduction which allows a thorough understanding of the concepts of "free energy", quantitation of energy available from ATP, and quantitation of strength of reducing or oxidising reactions. These latter aspects should only be reviewed if you have a background in Biochemistry (it is too detailed for students without that background).
An excellent description of oxidative phosphorylation can be found in this textbook of molecular cell biology, but it is excessively lengthy. It might be useful if you cannot gain access to Lehninger et al and need greater detail than that of Morrison et al.
This text is comprehensive but will be excessively detailed for most students. Chapters 3 and 15 are very useful in introducing prior concepts of free energy and oxidation-reduction reactions. The treatment is sufficiently quantitative and detailed to challenge those with prior undergraduate degrees in Biochemistry.
In case you wish to consider this topic before you secure access to any textbook, some annotated minimalist diagrams are included in the Appendix. The coloured and detailed diagrams in the textbooks, together with the text at the level appropriate to your background, are much better sources for learning than is the Appendix, but it might be useful as an "aide memoir" and to get you started if textbooks are difficult to secure.
Oxidation is the loss of electrons, whether to oxygen or to
some other molecule. Reduction is the gain of electrons. In biological
systems there are four types of electron transfer:
Oxidative phosphorylation can be understood qualitatively, which might satisfy most without previous Biochemistry. For students who have previous Biochemistry, you should revise the quantitative aspects of reduction potentials, for which it is necessary to understand units for free energy change and for reduction potential.
It is convenient to biochemists to adopt conventions which
define the strength of a reducing agent in terms of its potential
to donate electrons, and of an oxidising agent in terms of its
potential to accept electrons, in each case relative to the reduction
potential of the H+:H2 couple which is defined
as 0 volts at pH 7. The units of the standard reduction potential
are volts, the symbol is E'o; a strong reducing agent (such
as NADH) is poised to donate electrons and has a negative reduction
potential, whereas a strong oxidising agent (such as O2)
is ready to accept electrons and has a positive reduction potential.
Thus reduction potentials measure affinity for electrons.
[The theory of reduction potentials is well explained by Lehninger
et al, pp 385-388.]
Test your understanding of this concept by arranging the standard reduction potentials of the following biologically-important half reactions in a sequence, starting with strong electron donors (strong reducing agents) and ending with good electron acceptors (which are strong oxidising agents). Compare this order with that of the mitochondrial electron transport chain which you will read/have read about below.
| Half reaction | E'o (Volts) |
|---|---|
| Cytochrome a (Fe3+) + e- --> (Cytochrome a (Fe2+) | 0.29 |
| Cytochrome b (Fe3+) + e- --> Cytochrome b (Fe2+) | 0.077 |
| Cytochrome c (Fe3+) + e- --> Cytochrome c(Fe2+) | 0.254 |
| Cytochrome c1 (Fe3+) + e- --> Cytochrome c1(Fe2+) | 0.22 |
| FAD + 2H+ + 2e- --> FADH2 | -0.219 |
| Fe3+ + e- --> Fe2+ | 0.771 |
| 2H+ + 2e- --> H2 (at standard conditions, pH 0) | 0.00 |
| 2H+ + 2e- --> H2 (at pH 7, biochemists'standard) | -0.414 |
| NAD+ + H+ + 2e- --> NADH | -0.320 |
| NADP+ + H+ + 2e- --> NADPH | -0.324 |
| 1/2 O2 + 2H+ + 2e- --> H2O | 0.816 |
| Ubiquinone + 2H+ + 2e- --> Ubiquinol + H2 | 0.045 |
Table 1. Selected reactions and their reduction potentials
It is also convenient to biochemists to define the standard
free-energy change of a reaction at pH 7: the units are kJ/mol
and the symbol is Go'. [Note: the delta sympbol
() may not appear correctly on some browsers-it symbolises a change
in Go'] The change in free energy of a reaction
is a criterion of whether the reaction can occur spontaneously:
it can do so only if Go' is negative. The energy
available from the cleavage of different chemical bonds can be
compared, by comparison of the Go' values for
each reaction.
[The theory underlying this term can be obtained on p. 189 of
Dow et al, or more clearly but in more detail in pp. 364-381 of
Lehninger, Nelson & Cox.]
All students should be confident in answering the review questions, and be able to attempt the applied questions 1, 2, 3 and 5. The applied questions 4 and 6 should be attempted only if you have prior biochemistry.
Many texts contain very pertinent questions which you could access; some are gathered below.
What conditions might lead to depletion of the supply of (a) oxygen or of (b) NADH?
Calculate the rate at which this tissue consumes glucose and produces ATP. If the steady-state concentration of ATP in rat heart muscle is 5 micromol/g tissue, calculate the time required, in seconds, to completely turn over the cellular pool of ATP.
What does this result indicate about the need for tight regulation of ATP production? Would you expect these data for rat heart to be approximately or exactly representative of the corresponding results for human heart, or quite dissimilar from the energy requirements of human heart, after allowing for size differences?
What does this observation tell you about the metabolic requirements of the optic nerve?
Oxidative phosphorylation is the process in which ATP is formed as the electrons are transferred from NADH or FADH2 to O2 by a series of electron carriers.
(Go' = -52.6 kcal/mol)
The cytochromes are small proteins, each carrying a haem group with a single atom of iron. They are thus able to carry only one electron.
Fe3+ + e- --> Fe2+
The heme in cytochromes c and c1 is covalently attached to two cysteine side chains, as illustrated in the diagram below.
Three dimensional structure of reduced Cytochrome c from Tuna.
The heme group is shown in red, and Fe in purple. The two structures shown (above) are of cytochrome c; the wireframe on the left and a backbone model (which shows the peptide bond backbone of the protein).
All four complexes are located in the inner mitochondrial membrane. The arrangement of the complexes is illustrated diagrammatically next.
The free energy change at each step of the electron transport chain is shown on the following graph:
The favourable redox reactions which occur during the flow of electrons from NADH to O2 are "coupled" to the synthesis of ATP.
The energy released upon oxidation of NADH is trapped initially as a proton gradient across the inner mitochondrial membrane: this gradient is used by an enzyme (ATP synthase) to drive the synthesis of ATP.
Thus, all the above processes are rate limited by the slowest step; this is called "respiratory control" e.g. if O2 supply is cut off all the other processes stop.
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