5.7.3 – Explain how phosphorylation of ATP requires energy and how dephosphorylation of ATP provides an immediate supply of energy for biological processes


Adenosine TriPhosphate (ATP) is made from three components;


  • Ribose (the same sugar that forms the basis of DNA).
  • A base (a group consisting of linked rings of carbon and nitrogen atoms); in this case the base is adenine.
  • Up to 3 phosphate These phosphates are the key to the activity of ATP

The energy used in all cellular reactions comes from ATP. By breaking the 3rd phosphate from the ATP molecule energy is released, which can be used to power intracellular reactions. The ATP is then regenerated by recombining the phosphate and ADP in respiration (or another process e.g. photosynthesis).


The recycling of ATP is crucial for life. For example a runner uses ~84kg of ATP in a marathon (more than their total body weight), yet there are only 50g of ATP     in the


entire body! This means each that each molecule of ATP has been recycled 1676 times during the race!


Normally, as soon as ATP has been converted into ADP + Pi it is converted back into ATP using energy from respiration. However, during exercise ADP may be converted into AMP or even Adenosine to provide energy.


Respiration: a process in which the chemical bond energy in glucose molecules is used to convert 38 ADP molecules into 38 ATP molecules. Oxygen is required and Carbon Dioxide and Water are produced as waste products.

Respiration occurs in 4 distinct steps;


Glycolysis takes place in the cytoplasm of a cell

In Glycolysis a Glucose molecule (6C) is split into 2 molecules of Glyceraldehyde Phosphate (3C).  2ATPs are required for this to happen.

Then, each 3C Glyceraldehyde Phosphate molecule is converted into a 3C Pyruvate molecule. In the process of converting one Glyceraldehyde Phosphate to one Pyruvate, enough energy is released to convert one NAD molecules into one NADH molecules and also to make two ATP molecules.

Overall; 4ATP are made, 2NADH are made and 2ATPs are used.

Net gain: 2ATP and 2NADH

In anaerobic conditions [H+] rises in the mitochondria as there are no available oxygen molecules to mop it up with and form water. This leads to saturation of the electron transport chain and a build-up of NADH and FADH2. This means [NAD] falls, which stops the Krebs’ Cycle. Acetyl CoA levels build-up, [CoA] falls and the Link Reaction stops. Pyruvate levels start to rise…

Muscle cells turn pyruvate into lactate to stop rising [pyruvate] from stopping Glycolysis (remember, enzyme controlled reactions are reversible and depend on [reactants] and [products]).

In the liver the lactate is converted back into pyruvate. This requires oxygen, which is the basis of the “Oxygen Debt”



Link Reaction takes place in the matrix of the mitochondria

In the Link Reaction a Pyruvate molecule (3C) is split into a 2C molecule and a CO2. The 2C molecule is attached to a CoA enzyme, forming Acteyl CoA.

Remember, two molecules of Pyruvate were made at the end of Glycolysis,  therefore the Link Reaction happens twice.

Overall; 2NADH and 2 CO2 are made. Net gain: 2NADH


2 NADH are made (4 overall)                  1 ATP is made (2 overall)

1 FADH2  is made (2 overall)                             2 CO2  are made (4 overall)

Krebs’ Cycle takes place in the matrix of the mitochondria

In the Krebs’ Cycle the Acetyl CoA gives its 2C atoms to a 4C molecule (Oxaloacetate) forming an unstable 6C molecule (Citric Acid). The 6C molecule breaks down into a  4C compound (Succinyl – CoA) releasing enough energy to make one NADH. The two spare C atoms are released as two CO2 molecules.

Succinyl – CoA is converted back into Oxaloacetate and this releases enough energy to make one NADH, one FADH2 and one ATP. The Oxaloacetate can then be used in the cycle again.

Remember, two molecules of Acetyl CoA were made at the end of the Link Reaction, therefore the Krebs’ Cycle happens twice.

Overall; 4NADH, 2FADH2, 2CO2  and 2ATP are made.



Oxidative Phosphorylation uses the NADH and FADH2 produced in the previous steps of respiration to make ATP. Each NADH makes 3ATP and each FADH2 makes 2 ATP.

Oxidative Phosphorylation takes place using enzymes embedded in the inner membrane of cristae of the mitochondria

Hydrogen atoms from the NADH and the reduced FADH2 are passed onto 2 the first 2 enzymes of the Electron Transport Chain. These enzymes are Hydrogen Carriers and they accept the H atoms from the NADH and the FADH2.

Electrons, which made up the chemical bond between the hydrogen atoms and the NADH / FADH2 are passed onto 3 Electron Carrier enzymes further down  the  Electron Transport Chain.

At the end of the Electron Transport Chain, the electrons are recombined with the H+ atoms and oxygen, to form water. This is the only, but crucial, part of respiration to involve oxygen.

NADH starts at the first Hydrogen Carrier and has enough energy to phosphorylate 3ADP. FADH2 has less energy and starts at the second Hydrogen Carrier, it generates 2 ATPs

Where does the 38 ATP come from?


Glycolysis produces;


Link Reaction produces;




Kreb’s Cycle produces;2ATP6NADH2 FADH2


Each NADH produces 3ATP \ total production is 30ATP from NADH

Each FADH2 produces 2ATP \ total production is 4ATP from FADH2

Grand Total         4ATP     +      30ATP  +       4ATP    = 38ATP

Chemiosmosis of H+ ions from the mitochondrial envelope into the matrix through ATP Synthetase proteins is what actually generates the ATP in respiration.




The electron transport chain uses the process of chemiosmosis (the diffusion of ions across a membrane). H+ ions are actively pumped into the mitochondrial envelope. This is done by the proteins in the electron transport chain, using the energy stored in NADH and FADH2.

The [H+] builds up to very high levels in the envelope. However, H+ cannot escape because it is charged (hydrophilic) and therefore cannot move through the phospholipid bilayer in the envelope membranes.

Special proteins called ATP Synthetase do allow H+ to pass through them and escape into the mitochondrial matrix. Whenever an H+ ion moves through the ATP Synthetase protein an ADP is phosphorylated by the ATP Synthetase.


In summary;


  1. NADH and FADH2 contain stored chemical energy.
  2. The energy is used to pump H+ into the mitochondrial membrane against the concentration
  3. H+ trapped in one place represents a store of potential
  4. H+ ions leave the envelope through ATP Synthetase
  5. The potential energy of the H+ is used to phosphorylate ATP as the H+ moves out of the envelope.