Oxidative phosphorylation

1. PRESENTATION OF BIOCHEMISTRY TOPIC: Oxidative Phosphorylation

2. RESPIRATION Respiration – a process by which cells derive energy with a controlled reaction between H + and O 2 ; the end product being water. Aerobic organisms are able to capture a far greater proportion of the available free energy of respiratory substrates than anaerobic organisms

3. RESPIRATION The objective of respiration is to produce ATP. Energy is released from oxidation reactions in the form of electrons Electrons are shuttled by electron carriers (e.g. NAD+) to an electron transport chain Electron energy is converted to ATP in the electron transport chain

4. METABOLISM Metabolism is the sum of the chemical reactions in an organism. Catabolism is the energy-releasing processes. Anabolism is the energy-using processes. Catabolism provides the building blocks and energy for anabolism.

5. ATP couples energy between catabolism and anabolism Energy available for work & chemical synthesis (e.g. movement, signal amplification, etc . A nabolism ATP ADP + P i Energy from food (fuel molecules) or from photosynthesis Catabolism

6. Oxidative phosphorylation is the process by which the energy stored in NADH and FADH2 is used to produce ATP. A. Oxidation step: electron transport chain NADH + H + + O 2 NAD + + H 2 O FADH 2 + O 2 FAD + H 2 O B. Phosphorylation step ADP + Pi ATP Oxidative Phosphorylation

7. Mitochondria, have been termed the "powerhouses" of the cell since the final energy release takes place in the mitochondria only. Mitochondria have an outer membrane that is permeable to most metabolites, an inner membrane that is selectively permeable, enclosing a matrix within . MITOCHONDRIA

8. The outer membrane is characterized by the presence of various enzymes, including acyl-CoA synthetase and glycerol phosphate dehydrogenase. Adenylyl kinase and creatine kinase are found in the intermembrane space. The phospholipid cardiolipin is concentrated in the inner membrane together with the enzymes of the respiratory chain, ATP synthase and various membrane transporters. The matrix encloses the enzymes of TCA cycle, beta oxidation and pyruvate dehydrogenase complex. MITOCHONDRIA

9. MITOCHONDRIA

10. The electron transport chain is series of protein complexes embedded in mitochondrial membrane . This chain is consist of 4 complexes and ATP Synthesis. 4 complexes are : NADH FADH C ytochrome b-c Cytochrome oxidase ELECTRON TRANSPORT CHAIN

11. ELECTRON TRANSPORT CHAIN E lectrons are come from electron carriers and they travel through the electron transport chain where the electrons final destination is oxygen which will help to reduce Oxygen to form water. So oxygen is known as final acceptor. Electrons captured from donor molecules are transferred through 4 complexes.

12. (Complex 1 ) NADH-coenzyme Q oxidoreductase , also known as NADH dehydrogenase or complex I, is the first protein in the electron transport chain NADH

13. NADH In Complex I, In which NADH dehydrogenase oxidized to NAD+H+,this process obtain two electrons which will first given to FMN( flavin mononucleotide) from here the electrons are transfered one at a time through a series of iron sulfur center than 2 electrons create a proton gradient which bring 2 hydrogen ions from the matrix and bound to ubiquinone and as a biase ubiquinone it will reduced to ubiquinol (QH2 ). Complex I can transfer 4 protons from the matrix inner membrane space.It will be seen that transfer of four protons in to inner membrane space is equivalent to formation of one ATP molecule.

14. FADH 2 ( C omplex 2 ) Succinate-Q oxidoreductase , also known as complex 2 or succinate dehydrogenase,(from the citric acid cycle)is a second entry point to the electron transport chain .

15. FADH 2 It contains FAD( Flavin adenine dinucleotide) and Fe-S centers; it lacks proton pump activity. It oxidizes succinate to fumarate and reduces ubiquinone. The two hydrogen atoms are first taken up by FAD to form FADH2 then passed through a series of iron sulfur centers and passed to ubquinone . As this reaction releases less energy than the oxidation of NADH, complex II does not transport protons across the membrane and does not contribute to the proton gradient.

16. FADH 2 For this reason, whereas transfer of two hydrogen from NADH+H+ to coenzyme Q by the complex 1 results in formation of one ATP,the transfer of two H atoms from FADH2 to coenzyme Q does not give rise to any ATP.

17. Coenzymes Q ubiquinone flows to the inner membrane its purpose is to carry electron through different complexes because it is a mobile protein where the complexes are stationary coenzymes travel to the inner membrane with 2 electrons.It would not associate with complex 2 but it would associate with complex 3. Coenzymes Q ubiquinone

18. Complex 3 Cytochrome b-c complex also called cytochrome c oxireductase Complex 3 has a few important sub unit or 3 imp structure Iron sulphr (Fe-S) protein Cytochrome b Cytochrome c C ytochrome b-c

19. Cytochromes are protein containing heme group. When Electrons are donated from NADH to NADH dehydrogenase, a large protein complex that pumps protons across the inner membrane . Then, electrons are transported to the coenzyme Q (Q), also termed ubiquinon ; then ubiquinon travel to the inner membrane and associate with the subunit of complex 3. Cytochrome b-c

20. Cytochrome b-c In complex 3 cytochrome c is not a part of any enzyme complex, is freely soluble and occurs in the inter membrane space Cytochrome c is also called mobile protein because it travel to the inter membrane space and attached or bind to the complex 4 cytochrome oxidase.

21. Complex 4 The final step of ETC is the reduction of molecular oxygen by electrons derived from cyt -c. Complex 4 consist of 3 important sub unit Subunit 1 has two heme group a and a 3 Subunit 2 contains two Cu ions Subunit 3 is essential for the activity of complex 4 Cytochrome oxidase

22. The cytochrome oxidase complexes then transfer electrons from cytochrome c to oxygen, the terminal electron acceptor, and water is formed as the product . Cytochrome oxidase also pumps 2 protons across the membrane . The transfer of protons generates a proton motive force across the membrane of the mitochondrion. Cytochrome oxidase

23. Cytochrome oxidase Electrons are transported between all these complexes and where will rise at oxygen so oxygen is final electron aceptor . These electrons are come from 1NADH and so now if we calculate all the protons pumped from 1 NADH to all the complexs . Complex 1 = 4 protons Complex 3 = 4 protons Complex 4 = 2 protons These 10 hydrogen ion it would go through the ATP synthase to produce ATP

24. Chemiosmotic Theory The chemiosmotic theory was developed by the British biochemist, Peter Mitchell which explain the mechanism of ATP formation. According to this theory, the tranfer of electrons down an electron transport system through a series of oxidation-reduction reactions releases energy .As electrons are transferred along the electron Transport chain from electron donor to electron acceptor in the inner mitochondrial membrane,free energy is released. This energy allows certain carriers in the chain to transport hydrogen ions (protons) which thus contains a higher concentration of protons than the matrix. This creates an electrochemical gradient across the inner membrane. The energized state of the membrane as a result of this charge separation is called proton motive force or PMF.

25. Chemiosmotic Theory This proton motive force provides the energy necessary for enzymes called ATP synthases, to catalyze the synthesis of ATP from ADP and phosphate. This generation of ATP occurs as the protons across the membrane through the ATP synthase complexes re-enter the matrix of the mitochondria. As the protons move down the concentration gradient through the ATP synthase, the energy released causes the rotor (F0) and stalk of the ATP synthase to rotate. The mechanical energy from this rotation is converted into chemical energy as phosphate is added to ADP to form ATP in the catalytic head (F1 domain)

26. The Generation of ATP ATP is generated by the phosphorylation of ADP

27. ATP SYNTHASE COMPLEX The ATP synthase has two distinct subunits: T he transmembrane   F0 subunit , which contains a protein channel for the flow of protons . T he   F1 subunit , which protrudes into the matrix space and catalyzes the synthesis of ATP from ADP and inorganic phosphate

28. ATP synthase is embedded in the inner membrane, together with the respiratory chain complexes . Several subunits of the protein form a ball-like shape arranged around an axis known as F 1 , which projects into the matrix and contains the phosphorylation mechanism . F 1 is attached to a membrane protein complex known as F , which also consists of several protein subunits. F spans the membrane and forms a proton channel . ATP SYNTHASE COMPLEX

29. ATP SYNTHASE COMPLEX The flow of protons through F causes it to rotate, driving the production of ATP in the F 1 complex . A portion of the F1 subunit termed the stalk links the two subunits . As protons flow through the channel in the F0 subunit, they cause the embedded stalk to rotate in the stationary F1 subunit, thereby converting the energy of the electrochemical gradient into mechanical energy.

30. ATP SYNTHASE COMPLEX As the stalk rotates in one direction, it induces conformational changes in the proteins of the F1 subunit, which, in turn, catalyze the synthesis of ATP - thereby converting the mechanical energy of stalk rotation to chemical bond energy. Approximately 4 protons must pass through the ATP synthase complex for one ATP molecule to be synthesized

31. ATP SYNTHASE COMPLEX The hydrogen concentration is much greater in the inter membrane space than in the matrix, thus generating an electrochemical proton gradient. This gradient drives protons back across the inner membrane through the ATP synthase (shown in gray) that catalyzes the synthesis of ATP from ADP and inorganic phosphate (Pi).