respiration

• Mitochondria

  • Have two phospholipid membranes
  • Outer membrane is smooth and permeable to several small molecules
  • Inner membrane is folded (cristae) and less permeable
  • Site of the electron transport chain (used in oxidative phosphorylation)
  • Location of ATP synthase (used in oxidative phosphorylation)
  • Intermembrane space has a low pH due to the high concentration of protons
  • Concentration gradient across the inner membrane is formed during oxidative phosphorylation and is essential for ATP synthesis
  • Matrix is an aqueous solution within the inner membranes containing ribosomes, enzymes and circular mitochondrial DNA necessary for mitochondria to function
• structure of mitochondria

  A) matrix

  B) granule

  C) mitochondrial DNA

  D) ribosome

  E) outer membrane

  F) inner membrane

  G) cristae

  H) intermembrane space

  
• glucose + oxygen →  carbon dioxide + water + energy
• • Autotrophs are organisms that are able to synthesise their own usable carbon compounds from carbon dioxide in the atmosphere through photosynthesis
• • Heterotrophs: require a supply of pre-made usable carbon compounds which they get from their food
• • Aerobic respiration is the process of breaking down a respiratory substrate in order to produce ATP using oxygen
• • The process of aerobic respiration using glucose can be split into four stages:
  • Glycolysis takes place in the cell cytoplasm
  • The Link reaction takes place in the matrix of the mitochondria
  • The Krebs cycle takes place in the matrix of the mitochondria
  • Oxidative phosphorylation occurs at the inner membrane of the mitochondria
•  1. Glycolysis: Phosphorylation and splitting of glucose

  2. Link reaction: Decarboxylation and dehydrogenation of pyruvate 

  3. Krebs cycle: Cyclical pathway with enzyme-controlled reactions 

  4. Oxidative phosphorylation: Production of ATP through oxidation of hydrogen atoms
• Aerobic Respiration

  Glycolysis
• Glycolysis

  1. Trapping glucose in the cell by phosphorylating the molecule

  2. Splitting the glucose molecule in two

  3. Production of 2 Pyruvate (3C) molecules

  4. Net gain 2 ATP

  5. 2 reduced NAD
• Steps of glycolysis

  1. Phosphorylation: glucose (6C) is phosphorylated by 2 ATP to form fructose bisphosphate (6C)

  2. Lysis: fructose bisphosphate (6C) splits into two molecules of triose phosphate (3C)

  3. Oxidation: hydrogen is removed from each molecule of triose phosphate and transferred to coenzyme NAD to form 2 reduced NAD

  4. Dephosphorylation: phosphates are transferred from the intermediate substrate molecules to form 4 ATP through substrate-linked phosphorylation

  5. Pyruvate is produced: the end product of glycolysis which can be used in the next stage of respiration
• process of glycolysis

  A) glucose

  B) phosphorylation

  C) bisphosphate

  D) triose phosphate

  E) triose phosphate

  F) oxidation

  G) dephosphorylation

  H) pyruvate

  I) pyruvate

  
• glycolysis results in the production of:

  • 2 Pyruvate (3C) molecules
  • Net gain 2 ATP
  • 2 reduced NAD
• link reaction:

  • The end product of glycolysis is pyruvate
  • Pyruvate contains a substantial amount of chemical energy that can be further utilised in respiration to produce more ATP
  • When oxygen is available pyruvate will enter the mitochondrial matrix and aerobic respiration will continue
  • Pyruvate moves across the double membrane of the mitochondria via active transport
  • It requires a transport protein and a small amount of ATP
  • Once in the mitochondrial matrix pyruvate takes part in the link reaction
• The Link Reaction

  • The link reaction takes place in the matrix of the mitochondria
  • It is referred to as the link reaction because it links glycolysis to the Krebs cycle
  • The steps are:
  • Pyruvate is oxidised by enzymes to produce acetate, CH3CO(O)-  and carbon dioxide, requiring the reduction of NAD to NADH
  • Combination with coenzyme A to form acetyl coenzyme A (acetyl CoA)
  • It produces:
  • Acetyl coA
  • Carbon dioxide (CO2)
  • Reduced NAD (NADH)

  pyruvate + NAD + CoA → acetyl CoA + carbon dioxide + reduced NAD
• link reaction which occurs in mitochondrial matrix

  A) pyruvate

  B) acetyl coA

  C) coA

  
• Coenzyme


  A molecule that helps an enzyme carry out its function but is not used in the reaction itself
• Coenzyme A

  • Consists of a nucleoside (ribose and adenine) and a vitamin
  • Binds to the remainder of the pyruvate molecule (acetyl group 2C) to form acetyl CoA
  • Supplies the acetyl group to the Krebs cycle where it is used to continue aerobic respiration
• Role of coenzyme A

  1. Brings part of the carbohydrate (or lipid/amino acid) into the further stages of respiration

  2. Links the initial stage of respiration in the cytoplasm to the later stages in the mitochondria
• Krebs Cycle

  consists of a series of enzyme-controlled reactions
• • 2 carbon (2C) Acetyl CoA enters the circular pathway from the link reaction in glucose metabolism
  • Acetyl CoA formed from fatty acids (after the breakdown of lipids) and amino acids enters directly into the Krebs Cycle from other metabolic pathways
  • 4 carbon (4C) oxaloacetate accepts the 2C acetyl fragment from acetyl CoA to form the 6 carbon (6C) citrate
  • Coenzyme A is released in this reaction
  • Citrate is then converted back to oxaloacetate through a series of oxidation-reduction (redox) reactions
• Kreb's cycle

  A) 6C: citrate

  B) 5C: oxyacetate

  
• Regeneration of Oxaloacetate

  • Oxaloacetate is regenerated in the Krebs cycle through a series of redox reactions
  • Decarboxylation of citrate
  • Releasing 2 CO2 as waste gas
  • Oxidation (dehydrogenation) of citrate
  • Releasing H atoms that reduce coenzymes NAD and FAD
  • 3 NAD and 1 FAD → 3NADH + H+ and 1 FADH2
  • Substrate-linked phosphorylation
  • A phosphate is transferred from one of the intermediates to ADP, forming 1 ATP
• • Coenzymes NAD and FAD play a critical role in aerobic respiration
  • Sources of reduced NAD & FAD
  • Reduced NAD:
  • 2 x 1 = 2 from Glycolysis
  • 2 x 1 = 2 from the Link Reaction
  • 2 x 3 = 6 from the Krebs cycle
  • Reduced FAD:
  • 2 x 1 = 2 from the Krebs cycle
• • When hydrogen atoms become available at different points during respiration NAD and FAD accept these hydrogen atoms
  • They transfer the hydrogen atoms (hydrogen ions and electrons) from the different stages of respiration to the electron transport chain on the inner mitochondrial membrane, the site where hydrogens are removed from the coenzymes
• • Oxidative phosphorylation is the last stage of aerobic respiration
  • It takes place at the inner mitochondrial membrane
  • It results in the production of many molecules of ATP and the production of water from oxygen
• current model for oxidative phosphorylation is the chemiosmotic theory
• Chemiosmotic theory

  • The model states that energy from electrons passed through a chain of proteins in the membrane (the electron transport chain) is used to pump protons (hydrogen ions) up their concentration gradient into the intermembrane space
  • The hydrogens are then allowed to flow by facilitated diffusion through a channel in ATP synthase into the matrix
  • The energy of the hydrogens flowing down their concentration gradient is harnessed (a bit like water flowing through a hydroelectric damn) resulting in the phosphorylation of ADP into ATP by ATP synthase
• Oxidative phosphorylation

  1. Hydrogen atoms donated by reduced NAD (NADH) and reduced FAD (FADH2) from the Krebs Cycle

  2. Hydrogen atoms split into protons (H+ ions) and electrons

  3. High energy electrons enter the electron transport chain and release energy as they move through

  4. Released energy used to transport protons across the inner mitochondrial membrane from the matrix into the intermembrane space

  5. Concentration gradient of protons established between the intermembrane space and the matrix

  6. Protons return to the matrix via facilitated diffusion through the channel protein ATP synthase

  7. Movement of protons down their concentration gradient provides energy for ATP synthesis

  8. Oxygen acts as the 'final electron acceptor' and combines with protons and electrons at the end of the electron transport chain to form water
• The electron transport chain

  • The electron transport chain is made up of a series of membrane proteins/ electron carriers
  • They are positioned close together which allows the electrons to pass from carrier to carrier
  • The inner membrane of the mitochondria is impermeable to hydrogen ions so these electron carriers are required to pump the protons across the membrane to establish the concentration gradient
• oxidative phosphorylation via chemiosmotic theory occurs on inner mitochondrial membrane and requires NADH and FADH2

  
• Oxygen acts as the final electron acceptor.
• Anaerobic pathways

  • Some cells are able to oxidise the reduced NAD produced during glycolysis so it can be used for further hydrogen transport
  • This means that glycolysis can continue and small amounts of ATP are still produced
  • Different cells use different pathways to achieve this
  • Yeast and microorganisms use ethanol fermentation
  • Other microorganisms and mammalian muscle cells use lactate fermentation
• Ethanol fermentation

  • In this pathway reduced NAD transfers its hydrogens to ethanal to form ethanol
  • In the first step of the pathway pyruvate is decarboxylated to ethanal
  • Producing CO2
  • Then ethanal is reduced to ethanol by the enzyme alcohol dehydrogenase
  • Ethanal is the hydrogen acceptor
  • Ethanol cannot be further metabolised; it is a waste product

  A) ADP

  B) glucose

  C) pyruvate

  D) ethanal

  E) ethanol

  
• Lactate fermentation

  • In this pathway reduced NAD transfers its hydrogens to pyruvate to form lactate
  • Pyruvate is reduced to lactate by enzyme lactate dehydrogenase
  • Pyruvate is the hydrogen acceptor
  • The final product lactate can be further metabolised

  A) glucose

  B) ATP

  C) pyruvate

  D) lactate

  
• Metabolization of lactate

  • After lactate is produced two things can happen:

  1. It can be oxidised back to pyruvate which is then channelled into the Krebs cycle for ATP production
  2. It can be converted into glycogen for storage in the liver

  • The oxidation of lactate back to pyruvate needs extra oxygen
  • This extra oxygen is referred to as an oxygen debt
  • It explains why animals breathe deeper and faster after exercise
• Rate of respiration (sec-1) = 1 / time (sec)
• Apparatus

  • Yeast
  • Glucose solution
  • Test tubes
  • Stopwatch
  • DCPIP
• Method - Temperature

  1. Add a set volume of yeast suspension to test tubes containing a certain concentration of glucose

  2. Put the test tube in a temperature-controlled water bath and leave for 5 minutes to ensure the water temperature is correct and not continuing to increase or decrease

  3. Add a set volume of DCPIP to the test tube and start the stopwatch immediately

  4. Stop the stopwatch when the solution becomes colourless or lose all blue colour

  5. Record the time taken for a colour change to occur once the dye is added
• This is subjective and therefore the same person should be assigned this task for all repeat experiments