1. Introduction to metabolism

2. Metabolism Term used to describe all the chemical reactions occurring in an organism Metabolism = anabolism + catabolism Break down chemistry is called catabolism Buildup (synthesis) chemistry is called anabolism Most chemistry is assisted by proteins called enzymes

3. Sources of energy Carbohydrates, lipids, and proteins are sources of energy for the body Energy is stored in the electrons associated with C-H bonds Carbohydrates are the first source of energy in the body. Lipids contain the most of C-H bonds per gram and so have the highest number of Calories Energy from foods is converted to ATP: Cell energy

4. Building Materials Building materials may be consumed directly or synthesized from materials through metabolic pathway A metabolic pathway is a series of chemical steps that begin with a specific molecule and end with a product Each of the Chemical steps of the metabolic pathway is usually catalyzed by a specific enzyme. Enzyme 1Enzyme 2Enzyme 3 A B C D Reaction 1Reaction 2Reaction 3 Starting molecule Product

5. Catabolic pathways : Break down complex molecules into simpler compounds Release energy This energy is stored in organic molecules until need to do work in the cell. Anabolic pathways : Build complicated molecules from simpler ones Consume energy The energy released by catabolic pathways is used to drive anabolic pathways.

6. Bioenergetics Is the Study of how organisms manage their energy resources Energy : –- Is the capacity to cause change - Exists in various forms, of which some can perform work. Kinetic energy –Is the energy associated with motion Potential energy –- Is stored in the location of matter –- Includes chemical energy stored in molecular structure

7. ATP (adenosine triphosphate) O O O O CH 2 H OH H N HH O N C HC N C C N NH 2 Adenine Ribose Phosphate groups O O O O O O - --- CH Immediate source of cellular energy Common to ALL living things Made of adenine, ribose and three phosphates Made by each cell 10 7 molecules used and regenerated/second/cell

8. ATP An exergonic reaction –Proceeds with a net release of free energy and is spontaneous An endergonic reaction –Is one that absorbs free energy from its surroundings and is nonspontaneous ATP powers cellular work by coupling exergonic reactions to endergonic reactions Energy coupling Is a key feature in the way cells manage their energy resources.

9. ATP hydrolysis –Can be coupled to other reactions Endergonic reaction: ∆G is positive, reaction is not spontaneous ∆G = +3.4 kcal/mol Glu ∆G = - 7.3 kcal/mol ATP H2OH2O + + NH 3 ADP + NH 2 Glutamic acid Ammonia Glutamine Exergonic reaction: ∆ G is negative, reaction is spontaneous P Coupled reactions: Overall ∆G is negative; together, reactions are spontaneous ∆G = –3.9 kcal/mol

10. The Regeneration of ATP Catabolic pathways –Drive the regeneration of ATP from ADP and phosphate ATP synthesis from ADP + P i requires energy ATP ADP + P i Energy for cellular work (endergonic, energy- consuming processes) Energy from catabolism (exergonic, energy yielding processes) ATP hydrolysis to ADP + P i yields energy

11. Electron Transport Chain Food (glucose) is oxidized and the released hydrogens: –Are transported by coenzymes NADH and FADH 2 –Enter a chain of proteins bound to metal atoms (cofactors) –Combine with molecular oxygen to form water –Release energy The energy released is harnessed to attach inorganic phosphate groups (P i ) to ADP, making ATP by oxidative phosphorylation –“phosphorylation” - to add phosphate to a substance ADP + P ATP

12. Mechanism of Oxidative Phosphorylation The hydrogens delivered to the chain are split into protons (H + ) and electrons –The protons are pumped across the inner mitochondrial membrane to the intermembrane space –This creates a pH and concentration gradient (of H + ) –The electrons are shuttled from one acceptor to the next Electrons are delivered to oxygen, forming oxygen ions Oxygen ions attract H + that were pumped into the intermembrane space to form water H + that were pumped to the intermembrane space: –Diffuse down their gradients back to the matrix via ATP synthase (from greater to lesser concentration) –Release energy to make ATP

14. ATP Synthase The enzyme consists of three parts: a rotor, a knob, and a rod Current created by H + causes the rotor and rod to rotate This rotation activates catalytic sites in the knob where ADP and P i are combined to make ATP

15. 15 Mitochondrion Oxidative decarboxilation of pyruvate, & citric acid cycle take place in matrix, along with fatty acid oxidation Site of oxidative phosphorylation Permeable

16. Respiratory control: Rate of oxidative phosphorylation is determined by the need for ATP 1.Electron transport and ATP synthesis is coupled. 2. Electrons do not flow through the electron-transport chain to O2 unless ADP is simultaneously phosphorylated to ATP. 3. When [ADP] rises, the rate of oxid. phos. increases to meet the ATP needs. (This is the case in active muscle) 4. The regulation of the rate of oxid. phos. by the ADP level is called Respiratory control or Acceptor control.

17. Two Translocase Proteins 1.ATP/ADP Translocase –also called the adenine nucleotide translocase. –functions to export one ATP for every ADP that is imported. –an antiporter because it translocates molecules in opposite directions across the membrane. –for every ADP molecule that is imported from the cytosol, an ATP molecule is exported from the matrix. 2.Phosphate Translocase –translocates one P i and one H + into the matrix by an electroneutral import mechanism.

18.  Cyanide (CN - ), azide (N3 - ) react with the ferric (Fe 3+ ) form of heme a3.  Carbon monoxide (CO): Inhibits the ferrous (Fe 2+ ) form.  Inhibits electron flow from Cyt bH  Inhibits electron flow in Complex I  Prevents the utilization of NADH as a substrate. Site of action of some inhibitors of electron transport

19. Dinitrophenol is a hydrophobic molecule that remains in the mitochondrial membrane as a chemical uncoupler for a long time - a very dangerous way to burn fat. J Anal Toxicol. 2006 Apr;30(3):219-22.

20. Summary of known ETS and Ox Phos inhibitors You should be able to answer questions about changes in the rates of succinate oxidation, O2 consumption, and ATP synthesis in mitochondrial suspensions if provided information about any of these ETS, Ox Phos, or translocase inhibitors.

21. The UCP1 uncoupling protein, also called thermogenin, controls thermogenesis in newborn and hibernating animals Cell-specific expression of the UCP1 protein leads to heat production under aerobic conditions by short circuiting the proton gradient across the mitochondrial inner membrane. The UCP1 protein is expressed at high levels in special fat cells called brown adipose tissue which contain fatty acids for the production of acetyl CoA to drive NADH production by the citrate cycle, and large numbers of mitochondria to increase the output of heat by the electron transport system.

22. Problems Potent poisons: What is the effect of each of the following inhibitors on e transport and ATP formation by the respiratorty chain? a)Azide: It blocks e transport and proton pumping at Complex IV. b)Atractyloside: It block e transport and ATP synthesis by inhibiting the exchange of ATP and ADP across the inner mitochondrial membrane. c)Rotenone: It blocks e transport and proton pumping at Complex I. d)DNP: It blocks ATP synthesis without inhibiting e transport by dissipating the proton gradient. e)CO: It blocks e transport and proton pumping at Complex IV. f)Antimycin A: It blocks e transport and proton pumping at Complex III. Uncouplers of Oxidative Phosphorylation: In normal mitochondria the rate of e transfer is tightly coupled to the demand for ATP. Thus when ATP is demanded at a high rate, e transfer is rapid. Under such conditions of tight coupling, the no of ATP produced per atom of oxygen consumed when NADH is the e donor---know as the P/O ratio---is close to 3. a)increased activity of the respiratory chain in the presence of DNP requires the degradation of additional fuel. By oxidizing more fuel (including fat reserves) to produce the same amount of ATP, the body loses weight. When the P/O ratio approaches zero, the lack of ATP results in death.