Respiration (A-level only) — AQA A-Level Biology

Respiration transfers energy from organic molecules to ATP. ATP is synthesised by substrate-level phosphorylation in glycolysis and the Krebs cycle, and by oxidative phosphorylation. In oxidative phosphorylation, electrons from reduced coenzymes pass along the electron transfer chain, releasing energy to actively transport protons. The resulting proton gradient drives ATP synthase to phosphorylate ADP. ATP is the immediate energy source for cells. It is hydrolysed by ATP hydrolase to ADP and Pi, releasing manageable energy for processes such as active transport, muscle contraction and protein synthesis. Glycolysis initially requires ATP to phosphorylate glucose, so respiration both consumes and produces ATP. Phosphorylation makes glucose more reactive and less stable, allowing it to be split more readily; this is accepted as lowering the activation energy of the reaction.

Glycolysis happens in the cytoplasm of the cell, not inside the mitochondrion, and it needs no oxygen, which is what makes it the one stage common to aerobic and anaerobic respiration. Every organism carries it out, including prokaryotes, which have no mitochondria at all, and that universality is evidence of a shared evolutionary origin. Because glycolysis is anaerobic, a cell can keep making ATP when oxygen runs short; the limit is that the reduced NAD it produces must be re-oxidised, which anaerobically requires pyruvate to be converted to lactate or to ethanol. When oxygen is available, the pyruvate from glycolysis is moved into the mitochondrial matrix and oxidised further, so glycolysis is only the opening stage, worth a net two ATP out of the far larger yield a glucose molecule gives aerobically.

Glycolysis occurs in the cytoplasm. First, glucose is phosphorylated to glucose phosphate using ATP. This makes the molecule more reactive and lowers activation energy. The glucose phosphate is then split into two molecules of the three-carbon triose phosphate. Next, each triose phosphate is oxidised to pyruvate. During this oxidation, hydrogen is removed and accepted by NAD, forming reduced NAD. This stage also produces ATP via substrate-level phosphorylation. Because two ATP are used initially and four are produced, there is a net gain of two ATP and two reduced NAD per glucose molecule.

Without oxygen, the electron transfer chain stops and reduced NAD cannot be re-oxidised. Glycolysis would halt once cellular NAD is depleted. Anaerobic respiration solves this by oxidising reduced NAD, allowing glycolysis to continue. In animals, pyruvate acts as the hydrogen acceptor from reduced NAD, forming lactate. In plants and yeast, pyruvate is decarboxylated to ethanal, releasing carbon dioxide. Ethanal then accepts hydrogen from reduced NAD, forming ethanol. Both pathways regenerate oxidised NAD. The only ATP yield is the net two molecules from glycolysis. While often taught that lactate lowers pH and causes fatigue, this is a simplified theoretical model.

NAD is a coenzyme, and a cell holds only a small pool of it. Every triose phosphate oxidised in glycolysis loads one NAD with hydrogen, so unless that reduced NAD is unloaded again the pool is quickly exhausted and glycolysis stops, taking with it the cell's only anaerobic source of ATP. Aerobically, the unloading happens at the inner mitochondrial membrane, where reduced NAD is oxidised and passes its electrons to the electron transfer chain. Anaerobically, it happens when reduced NAD donates its hydrogen to pyruvate or to ethanal, forming lactate or ethanol. Either way, oxidised NAD returns to the cytoplasm and accepts hydrogen from the next triose phosphate, so glycolysis continues and the net two ATP per glucose keep being made.

Glycolysis occurs in the cytoplasm and produces pyruvate. For aerobic respiration, pyruvate must reach the mitochondrial matrix, where the link reaction and Krebs cycle occur. Pyruvate crosses the outer mitochondrial membrane, which is highly permeable, then crosses the inner mitochondrial membrane by active transport through a specific carrier protein. This transport requires energy and ensures a continuous supply of pyruvate for the link reaction. Glucose itself does not enter the mitochondrion because it is broken down to pyruvate in the cytoplasm during glycolysis. A common error is to state that glucose enters mitochondria; instead, pyruvate is the molecule transported into the matrix. The carrier protein is specific to pyruvate, and active transport allows pyruvate to move against its concentration gradient if necessary.

In the matrix, pyruvate is oxidised to acetate, producing reduced NAD and releasing carbon dioxide. Acetate combines with coenzyme A to form acetylcoenzyme A. This reacts with a 4-carbon molecule to form a 6-carbon molecule, entering the Krebs cycle. Through redox reactions, it is decarboxylated and dehydrogenated back to the 4-carbon molecule, producing ATP by substrate-level phosphorylation and reduced coenzymes. These release electrons to the electron transfer chain. Energy released pumps protons across the inner membrane. Protons diffuse back via ATP synthase, driving ATP synthesis (chemiosmosis). Oxygen is the final electron acceptor, forming water. Breakdown products of lipids and amino acids can also enter the Krebs cycle as alternative respiratory substrates.

The organism is usually yeast, and the named variable might be temperature, substrate concentration or the type of sugar supplied. Rate can be measured by the volume of carbon dioxide produced, collected in a gas syringe, or by the time taken for a redox indicator such as methylene blue or TTC to change colour as it is reduced by hydrogen from the respiring cells. In a respirometer, carbon dioxide is absorbed by potassium hydroxide or soda lime, so the liquid movement in the manometer measures oxygen uptake alone, not carbon dioxide release. A control tube containing boiled yeast corrects for pressure changes caused by temperature. Controlled variables include the volume and concentration of the yeast suspension, the volume of substrate, pH, and the time allowed for equilibration in the water bath before readings begin.