BIOL2: Haemoglobin and Oxygen Dissociation Curves (RCl 8) 3. 2 BIOL2: Haemoglobin and Oxygen Dissociation Curves (RCl 8)   3.2.4 The variety of life is extensive and this is reflected in similarities and differences in its biochemical basis and cellular organisation.

Haemoglobin The haemoglobins are a group of chemically similar molecules found in many different organisms. Haemoglobin is a protein with a quaternary structure. The role of haemoglobin in the transport of oxygen. The loading, transport and unloading of oxygen in relation to the oxygen dissociation curve. The effects of carbon dioxide concentration.  

Candidates should be aware that different organisms possess different types of haemoglobin with different oxygen transporting properties. They should be able to relate these to the environment and way of life of the organism concerned.

Structure of haemoglobin. There is a group of haemoglobins, all chemically similar. Same general structure. All conjugated proteins. Primary structure = 4 polypeptide chains. Secondary structure = α helix. Tertiary structure = each chain folded into a precise shape – relates to function. Quaternary structure = 2 pairs of polypeptides. 2α and 2β chains. All 4 polypeptide chains are linked to from an almost spherical shape. Each have a prosthetic group, which is a haem group associated with it, which contains a ferrous (Fe2+) ion. Each Fe2+ ion can combine with a single oxygen molecule (O2). Process = oxygenation. In total 1 haemoglobin can combine with 4 O2 molecules.

The role of haemoglobin. = transport oxygen. To do this must: Readily associate with oxygen at surface where gaseous exchange occurs. Readily dissociate from oxygen at those tissues requiring it.

Clever as haemoglobin can change its affinity for oxygen under different condition. Achieves this by changing shape in the presence of carbon dioxide. Different haemoglobins have slightly different sequences of amino acids and therefore slightly different shapes. Depending on the shape, haemoglobin molecules range from those with a high affinity to those with a low affinity for oxygen. In presence of carbon dioxide haemoglobin binds more loosely to oxygen, so haemoglobin releases its oxygen more easily.

Process of haemoglobin combines with oxygen = loading or associating Process of haemoglobin combines with oxygen = loading or associating. Happens in lungs. Process of haemoglobin releases its oxygen = unloading or dissociating. Happens in tissues.

Carbon dioxide concentration Affinity of haemoglobin for oxygen Region of body Oxygen concentration Carbon dioxide concentration Affinity of haemoglobin for oxygen Result Gas exchange surface High Low Oxygen is attached Respiring tissues Oxygen is detached

Different haemoglobins Different organisms have different haemoglobin. They differ due to how they take up and release oxygen.  

Haemoglobins with a high affinity for oxygen. Take up oxygen more easily but release it less readily. E.g. of an organism that lives in an environment where there is little oxygen, so haemoglobin must be able to combine readily with oxygen if it is to absorb enough. Metabolic rate must not be too quick, and then it does not matter if oxygen is not released as readily into the tissues.

Haemoglobin with a low affinity for oxygen. Takes up oxygen less easily but release it more readily. E.g. organism with high metabolic rate needs to release oxygen readily into its tissues. As long as there is plenty of oxygen in the environment, then it is more important that the haemoglobin releases oxygen easily.

Questions Describe the quaternary structure of haemoglobin. 2. Explain how DNA leads to different haemoglobin molecules having a different affinity for oxygen. 3. When the body is at rest only 1 of the 4 oxygen molecules carried by haemoglobin is normally released into the tissues. Suggest why this could be an advantage when the organism becomes more active. 4. Carbon monoxide occurs in car exhaust fumes. It binds permanently to haemoglobin in preference to oxygen. Suggest a reason why a person breathing in car exhaust fumes might lose consciousness.  

Answers 1. 2 pairs of polypeptides, (2α and 2β) link to form a spherical molecule, (globular protein). Each polypeptide has a haem group that contains a ferrous ion. 2. Different base sequences in DNA- different amino acid sequences (different primary structure) – and so get different tertiary/quaternary structures and shape – different affinities for oxygen. 3. If all oxygen molecules were released there would be none in reserve to supply tissues when they are more active. 4. Carbon monoxide will gradually occupy all the sites on the haemoglobin instead of oxygen. No oxygen will be carried to tissues, such as the brain. Cells cease to respire and to function – person loses consciousness.

Oxygen dissociation curves. Measuring oxygen concentration Amount of gas in a mixture is measures by the pressure it contributes to the total pressure of the gas mixture = partial pressure of the gas. For oxygen written as pO2. For carbon dioxide = pCO2 Measured in kilopascals (kPa). The % of haemoglobin associated with oxygen at a given partial pressure of oxygen (pO2) = % saturation. Normal atmospheric pressure = 100kPa. Oxygen makes up 21% of atmosphere, so its partial pressure = 21kPa. In lungs partial pressure of oxygen = 13kPa and 98% of haemoglobin associates (binds) with oxygen. In respiring tissue at rest pO2 = 5.3kPa, with 73% haemoglobin associated with oxygen. In moderately respiring muscle pO2 = 2.5kPa, with 35% oxygen still associated with haemoglobin.

Draw an annotated graph here to show the % saturation of haemoglobin against the oxygen partial pressure.

HbO2 → Hb + O2 (reversible reaction) Oxyhaemoglobin Haemoglobin + Oxygen When haemoglobin is exposed to different partial pressure of oxygen it does not absorb oxygen evenly.

At very low concentrations of oxygen The 4 polypeptides of haemoglobin are tightly bound together. So difficult to absorb the 1st oxygen molecule, onto the first haem group. Once loaded this oxygen molecule causes the polypeptides load the 3 remaining oxygen molecules very easily, as the polypeptide chains open up, exposing the other 3 haem groups. This is why it is a sigmoid curve. Gradual increase in slope at beginning as 1st oxygen is difficult to join. 2 and 3 are easier and so steeper curve. The last part of the curve levels out, as the haemoglobin becomes saturated it gets harder for last oxygen to join. Where the curve is very steep a small change in pO2 causes a big change in the amount of oxygen carried by haemoglobin. Graph of this = oxygen dissociation curve

A small decrease in the partial pressure of oxygen leads to a lot of oxygen becoming dissociated from haemoglobin. Graph tails off at very high oxygen concentrations cos haemoglobin is almost saturated with oxygen. Are a large number of oxygen dissociation curves cos many types of haemoglobin and any 1 type of haemoglobin molecule can change under different conditions.

All have roughly the same shape but remember:   The further to the left the curve is – the greater the affinity of haemoglobin for oxygen, so it takes oxygen up result but releases it less easily. The further to the right the curve is – the lower the affinity of haemoglobin for oxygen, so it takes up oxygen less readily but releases it more easily.

Effects of carbon dioxide concentration. In the presence of carbon dioxide haemoglobin has a reduced affinity for oxygen. Haemoglobin gives up oxygen more readily at higher partial pressures of carbon dioxide. Cells respire and produce carbon dioxide. This raised the Pco2, so increase the rate of oxygen unloading and curve shifts down. The greater the conc of carbon dioxide the more readily the oxygen is released = the Bohr shift.

CO2 diffuses into red blood cells→ carbonic acid (H2CO3) enzyme = carbonic anhydrase. Carbonic acid dissociates → hydrogen ions and hydrogen carbonate ions. Most carbon dioxide in blood is carried in solution as hydrogen carbonate ions. The hydrogen ions are mopped up by haemoglobin → haemoglobinic acid.

Haemoglobin here is acting as a buffer, resisting the changes in blood pH when carbonic acid is formed. The formation of haemoglobinic acid forces haemoglobin to unload oxygen, causing the Bohr shift. This is why the behaviour of haemoglobin is different in the lungs than the tissues.

Lungs = gaseous exchange surface. Here low conc of carbon dioxide, so affinity of haemoglobin for oxygen is increased, which coupled with the high conc of oxygen means oxygen is readily loaded by haemoglobin. The reduced carbon dioxide level has shifted the oxygen dissociation curve to the left.

Respiring tissues E.g. muscles – with higher levels of CO2. The affinity of haemoglobin for oxygen is reduced. Added to the low concentration of oxygen in the muscles means oxygen is readily unloaded from haemoglobin into muscle cells. The increased CO2 has shifted the oxygen dissociation curve to the right.

Loading, transport and unloading of oxygen. The greater the concentration of carbon dioxide, the more readily haemoglobin releases its oxygen. Cos dissolved CO2 is acidic and the low pH causes haemoglobin to change shape.

At gas-exchange surface carbon dioxide is constantly removed. pH increases here due to low levels of CO2. High pH changes shape of haemoglobin so that oxygen is loaded more readily. This change in shape increases the affinity of haemoglobin for oxygen, so it is not released whilst being transported to the tissues. In tissue CO2 levels high due to respiration. Carbon dioxide dissolves in water to form carbonic acid. This reaction releases hydrogen ions So pH of blood drops. Lower pH changes shape of haemoglobin so it has a low affinity for oxygen. Haemoglobin releases oxygen into respiring tissues.

The higher the rate of respiration → the more CO2 tissues produce → the lower the pH → the greater the haemoglobin shape change → the more readily oxygen is unloaded → the more oxygen is available for respiration.

Humans – haemoglobin carries 4 oxygen molecules Humans – haemoglobin carries 4 oxygen molecules. Normally when resting only 1 of the 4 is unloaded at respiring tissues, so the haemoglobin that returns to the lungs still is 75% saturated. In an actively respiring tissue, then the 3 remaining oxygen molecules can be unloaded as well.

kPa 12 = Haemoglobins molecule is loaded with oxygen in the lungs. kPa 6 = Haemoglobin molecule in a resting tissue unloads 25% of its oxygen. kPa 2 = Haemoglobins molecule in an active tissue unloads 75% of its oxygen.

The partial pressure of oxygen at which haemoglobin is 95% saturated = the loading pressure. The partial pressure of oxygen at which haemoglobin is 50% saturated = the unloading pressure.

Different lives – Different haemoglobins.  High altitude adaptations in mammals. At high altitude – temp, humidity and pressure decreases. Oxygen partial pressure is lower, reducing the amount of taken up by blood. Can lead to inadequate amounts getting to respiring cells. If person goes up mountain slowly, then body adjusts = altitude acclimatisation. Get increase in haemoglobin content and increase in density of red blood cells in blood. With more haemoglobin the carrying capacity of haemoglobin increase but blood becomes thicker and requires more pressure to pump it around body.

Those that live at higher altitudes are born with higher red blood cell counts and have oxygen dissociation curves shifted to the left of a normal curve. Advantage cos it increases the oxygen saturation of haemoglobin at low oxygen partial pressure that occur at high altitude. Disadvantage – oxygen is unloaded less readily.

Foetal haemoglobin. Foetus in uterus gets oxygen by diffusion from mum’s placenta. Foetus has foetal haemoglobin which has a higher affinity for oxygen than maternal blood. So foetal haemoglobin has oxygen dissociation curve to left of maternal one.

Variations in amino acid sequences produce haemoglobin with different properties.

Lugworms. Live in the sand at the beach. Not very active, living in a U shape burrow. Is covered by sea water which circulates in its burrow. Oxygen diffuses into the lugworm’s blood, from the water and the haemoglobin transports it to the tissues respiring. The haemoglobin will be 90% saturated, even though partial pressure is so low. When the tide goes out it does not have fresh supply of oxygenated water. So water contains less and less oxygen, as lugworm uses it up.

Organisms with low conc of oxygen have haemoglobin with a high affinity for oxygen than human haemoglobin. The curve is to the left of the human one.

Questions 5. Explain why a lugworm can survive at these low concentrations of oxygen while a human cannot. 6. How is the lugworm able to obtain sufficient oxygen from an environment that contains so little? 7. Suggest 1 feature of a lugworm’s way of life that helps it to survive in an environment that has little oxygen. 8. Haemoglobin usually loads oxygen less readily when the conc of carbon dioxide is high, (the Bohr shift). The haemoglobin of lugworms does not exhibit this effect. Explain why to do so could be harmful. 9. Suggest a reason why lugworms are not found higher up the seashore. 10. Llamas live at high altitude. Here the atmospheric pressure is lower and so the partial pressure of oxygen is also lower. It is therefore difficult to load haemoglobin with oxygen. Suggest where the oxygen dissociation curve of llama haemoglobin is shifted to, relative to human haemoglobin.

Answers 5. At this partial pressure it is still 90% saturated. This is enough for a sedentary animal like the lugworm. For a human this low partial pressure would mean a much lower % saturation, more like 10%, not enough to keep cells alive. Haemoglobin has a high affinity for oxygen, so pick up oxygen easily and release it less readily. 6. The dissociation curve is shifted to the left. This means it is fully loaded with oxygen, even when there is little in the environment available. 7. Lugworm is not very active. So requires little oxygen. 8. Respiration produced carbon dioxide. This builds up in burrow. If lugworm exhibited the Bohr shift effect, it would not be able to absorb much oxygen when it was present in very low concentrations. 9. higher part of beach is uncovered for longer period of times, so lugworm would receive less frequent fresh sea water, during long times without fresh sea water, the lugworm would use up all oxygen and die. Higher up the beach, there will be drier sand and so the burrow will have less water in it and so less oxygen. 10. It is shifted to the left.

Diving mammals E.g. whales and seals. Would expect the, to have haemoglobin with high affinity for oxygen cos dive in deep water but not so, as take in air before they dive at the surface, so they don’t require high affinity for oxygen.

Small mammals Have a large surface area to volume ratio. So lose heat quickly. So to maintain temp have high metabolic rate to generate heat. Active = higher demand for oxygen, so have haemoglobin with a lower affinity for oxygen than human haemoglobin. So oxygen dissociation curve of a mouse is to the right of humans.

Questions 11. The oxygen dissociation curve of the mouse is shifted to the right of humans. What difference does this make to the way oxygen is unloaded from mouse haemoglobin compared to that of a human? 12. What advantage does this have for the maintenance of body temp in mice?

Answers 11. It unloads more readily. 12. Oxygen is more readily released from haemoglobin to the tissues. This helps tissue respire more and produce more heat, which helps maintain the body temp of a mouse.

Birds and fish. Flight in birds & swimming in fish both need energy. Flight muscles in wings need lots of oxygen to respire, to keep them airborne. So during flight they have a very high metabolic rate, to produce the energy to oppose gravity in air that gives little support. Fish have a different problem – they expend a lot of energy swimming cos water is very dense and difficult to move through.

Haemoglobin in root nodules Get haemoglobin in some plants and bacteria e.g. root nodules of leguminous plants like peas and beans. They have special haemoglobin like molecule = lepthaemoglobin. Nodules contain bacteria – they absorb nitrogen from air → ammonia = nitrogen fixation, using enzyme – nitrogenase. Must be in anaerobic conditions, but plant roots need aerated soil.

So leguminous plants have evolved the lepthaemoglobin from cytochrome molecules that are present in all cells. Lepthaemoglobin absorbs the oxygen in the root nodules and creates an oxygen free atmosphere for the nitrogen fixation bacteria.

Questions 13. Is the oxygen dissociation curve of a pigeon to the right or left of a human? Why? 14. Mackerel swim in the surface water of the sea. They swim fast to avoid predators. Plaice move slowly on the sea bed, camouflaged from predators. Both are approx same mass. Sketch a graph to show the positions of the oxygen dissociation curves for these 2 fish. 15. What is the effect of increased carbon dioxide concentration on oxygen dissociation? 16. How does this change the saturation of haemoglobin with oxygen? 17. A rise in temperature shifts the oxygen dissociation curve right. How does this enable exercising muscle to work more efficiently?

Answers 13. Shifted to the right so more oxygen is readily released to the tissues, so haemoglobin supplies more oxygen to respiring muscles. 14. Sigmoid curves. Plaice to the left of mackerel. 15. The curve is shifted to the right. 16. Haemoglobin has become less saturated. 17. Exercising muscle releases heat, shifting the curve to the right. This causes haemoglobin to release more oxygen for muscular activity and increased respiration.

Summary The whole story.