Wednesday,Jan 14 2009, 05:04:59 PMTransport of oxygen through blood- Myoglobin and..

Transport of oxygen through blood- Myoglobin and haemoglobin

Nazmul Islam

Faculty and Coordinator, Department of Chemistry GERF College (Out Reach post Graduate Centre), University of Mysore. Guest lecturer, Dept.of Chemistry, Chakda College, University of Kalyani. *Correspond to the author : nazmul.islam786@gmail.com/nazmul.islam@rediffmail.com

Terminology: The prefix haem (heme) comes from the Greek haima = blood. It turns up in haematite, iron(II) oxide, because of its red colour (Gk. haimatites = blood-like). Haemoglobin contains iron(II) - when oxidised it gives methaemoglobin containing iron(III). Similarly myoglobin is oxidised to metmyoglobin. The oxygenated forms are known as oxyhaemoglobin and deoxyhaemoglobin (different colour and structure), and oxymyoglobin and deoxymyoglobin. The free haem group contains iron(II), and when oxidised it gives haemin chloride. Dioxygen is used for clarity in the article to refer to the O2 molecule, to avoid confusion with oxygen atoms or oxygen as an element. Oxygen can occur in the body in several forms and it is reduced in stages from O.S. 0 to O.S. -2 in water, its most stable form. Intermediates include O2-, the superoxide ion, and O22-, the peroxide ion. Some facts and figures on blood The human body contains around 5L of blood. Each litre contains 5 x 1012 red blood cells and 150g of haemoglobin, equivalent to about 1.25 x 1021 haemoglobin molecules . The red cells are made in the bone marrow (2 million a sec.), a process controlled by the hormone erythropoietin (made in the kidneys). Each cell lasts around 120 days before they are broken up and their chemical components salvaged in the liver for reuse. Around 1% are replaced daily. Anaemia occurs when our blood contains too little haemoglobin. One teaspoon of blood should contain 5 billion red blood cells - when the concentration drops to around half the symptoms of anaemia develop. Figure 1 shows the relative sizes of a red blood cell, haemoglobin and a single haem unit. If we move to high altitudes, where the oxygen pressure is less, the body compensates by making extra red blood cells. (This is one reason why East African athletes who train at high altitudes do so well when running at sea level - they have more oxygen-carrying capacity in their blood). The production of red cells can be affected by drugs, by diseases like leukaemia, or shortage of vitamin B12 (a problem for some vegetarians), or a shortage of iron in the diet. Regular bleeding due to disease (stomach ulcers for example) or heavy menstrual periods in women, also results in a steady loss of the body's iron supply and the colour of the blood gets paler as anaemia sets in. Anaemia affects more women than men. The body should contain around 5g of iron - 65% is present in haemoglobin (in blood), 13% in ferritin (an iron store in the liver), 6% in myoglobin (in muscles) and the rest in the cytochromes and other molecules (16%). We cannot live without iron in our diet. Porphyrins, myoglobin and haemoglobin To understand oxygen transport in the body you must start with the structure of the relevant molecules. Both oxygen-carrying proteins - myoglobin (in muscles) and haemoglobin (in blood) contain the haem unit. This is a molecule based on a porphyrin ring structure, one which is widely used in biology as a complexing molecule for metal ions. It is a chelating ligand which attached itself to the metal ion at 4 or more points, thus producing very stable complexes (the chelate effect). Thus iron (and other metals) are soaked up by these complexing molecules and are very hard to remove again.. Difference between myoglobin and haemoglobin A hemoglobin molecule (GMM 64,500) consists of four polypeptide chains: two alpha chains, each with 141 amino acids and two beta chains, each with 146 amino acids. The protein portion of each of these chains is called "globin". The  and globin chains are very similar in structure. In this case,  and  refer to the two types of globin. Students often confuse this with the concept of  helix and  sheet secondary structures. But, in fact, both the  and  globin chains contain primarily  helix secondary structure with no  sheets. Figure 2 is a close up view of one of the heme groups of the human a chain from dexoyhemoglobin. In this view, the iron is coordinated by a histidine side chain from amino acid 87 (shown in green.) Each  or  globin chain folds into 8  helical segments (A-H) which, in turn, fold to form globular tertiary structures that look roughly like sub-microscopic kidney beans. The folded helices form a pocket that holds the working part of each chain, the heme. A heme group is a flat ring molecule containing carbon, nitrogen and hydrogen atoms, with a single Fe2+ ion at the center. Without the iron, the ring is called a porphyrin. In a heme molecule, the iron is held within the flat plane by four nitrogen ligands from the porphyrin ring. The iron ion makes a fifth bond to a histidine side chain from one of the helices that form the heme pocket. This fifth coordination bond is to histidine 87 in the human  chain and histidine 92 in the human  chain. Both histidine residues are part of the F helix in each globin chain. Figure 2 shows the porphyrin ring and the structure of the haem group, showing the coordination positions. Figure 3 compares the structures of myoglobin and haemoglobin. The oxygen is taken up by the iron(II) ion at the centre of the haem structure. It has one spare coordination position, which is usually occupied by a water molecule (a weak ligand). Dioxygen is a ã-bonding ligand that bonds more strongly than water to iron (but not as strongly as carbon monoxide). When the partial pressure of oxygen is high, water is displaced and oxygen is taken up, The way myoglobin and haemoglobin take up oxygen is different - a vital physiological fact. Figure 4 shows how the fraction of haem groups bonded to dioxygen varies with the partial pressure of oxygen in both myoglobin and haemoglobin. Remember that in the lungs the partial pressure of oxygen is high, whereas it is low in a muscle. In de-oxy Hb, Fe remains as high spin Fe(II). It is five coordinated and there is a possibility of H2O (solvent) molecule to loosely bonded in the sixth position. The structure is distorted octahedral. The HS size of Fe(t2g4 eg2) is not fit for the porphyrin ring whose diameter is near about 200nm. So this Fe remains 0.4 Å -0.6Å away from the central position and it may be at an angle of 80 with the perpendicular of the porphyrin ring. These form is called Tense form. When Hb comes at the lungs, then the pO2 is higher and Hb form oxy-Hb complex. H+Hb + O2=H+ + HbO2 When O2 coordinate at the sixth coordination position of Fe(II),it changes to low spin state (t2g configuration). The LS size of iron is smaller than HS size and it is fit for the grove of the porphyrin ring. Here the N atom of distal imidazole group form H bond with the coordinated oxygen and stabilized the oxyHb complex. This structure is called Relaxed structure of haemoglobin. This oxy Hb then moves to tissue or cell where CO2 forms due to metabolic action. In cell,pCO2 is higher, oxy Hb breakes. The CO2 reacts with H2O to form bicarbonate ion. CO2 +H2O H+ + HCO3- And here also pH is lower, so binding capacity of Hb is lower. But binding capacity of Mb is independent of pH. So, Mb form oxy complex. Mb+HbO2=MbO2+Hb [The Bohr Effect The ability of hemoglobin to release oxygen, is affected by pH, CO2 and by the differences in the oxygen-rich environment of the lungs and the oxygen-poor environment of the tissues. The pH in the tissues is considerably lower (more acidic) than in the lungs. Protons are generated from the reaction between carbon dioxide and water to form bicarbonate: CO2 + H20 -----------------> HCO3- + H+ This increased acidity serves a twofold purpose. First, protons lower the affinity of hemoglobin for oxygen, allowing easier release into the tissues. As all four oxygen’s are released, hemoglobin binds to two protons. This helps to maintain equilibrium towards the right side of the equation. This is known as the Bohr effect, and is vital in the removal of carbon dioxide as waste because CO2 is insoluble in the bloodstream. The bicarbonate ion is much more soluble, and can thereby be transported back to the lungs after being bound to hemoglobin. If hemoglobin couldn’t absorb the excess protons, the equilibrium would shift to the left, and carbon dioxide couldn’t be removed. In the lungs, this effect works in the reverse direction. In the presence of the high oxygen concentration in the lungs, the proton affinity decreases. As protons are shed, the reaction is driven to the left, and CO2 forms as an insoluble gas to be expelled from the lungs. The proton poor hemoglobin now has a greater affinity for oxygen, and the cycle continues. ] For de oxy Hb PKa value is higher than bicarbonate, so H+ of bicarbonate reacts to form H+Hb. As no single ion can move through blood, so HCO3- also move with H+Hb. The Hb is not a passive carrier of oxygen but it acts as integrated machine. It can be explain by comparing it with activity of Mb. Mb+O2 MbO2 Stability constant, KMb =[MbO2]/[Mb] PO2 If f be the fraction of Mb oxygenated, then KMb = [MbO2]/[Mb]PO2 = f/(1-f)PO2 Or, f = K PO2/(1+K PO2) Hb has four unit of heam group and four poly peptide chain, so if four heam group will be oxygenated at a time fHb= K PO24/(1+K PO24) but experimentally it is found that fHb= K PO2n/(1+K PO2n), n~2.8, and value depends upon pH. This can be explain by the theory of cooperativity. The haemoglobin curve is sigmoidial. When one unit is coordinate with O2, then the probability of oxygenation of the other unit increases,that is uptake of O2 molecules by one haem group of the haemoglobin makes uptake of O2 easier for the rest three This is called co operativity. i.e., K1 increased cellular O2 utilization -> decreased interstitual PO2 [15mmHg] ii) Venous blood: 100 ml blood combines with 4.4 ml O2 [20% sat; PO2 18 mmHg] iii) Thus, 15ml of O2 is transported by each 100 ml blood through the tissues per cycle iv) Therefore, increased cellular O2 utilization -> increase rate of O2 release from Hb • utilization coefficient i) utilization coefficient = fraction O2 released from blood as passes via tissue capillaries ii) normally 0.25 [25%] iii) strenuous exercise:- 0.75 - 0.85 • Hb helps maintain a constant PO2 in tissue fluids (oxygen buffer function of Hb) despite exercise or changes in atmospheric changes in PO2 • Effect of blood flow on metabolic use of oxygen i) total amount of O2 available each minute for use in any given tissue is determined by: a) quantity of O2 transported in each 100 ml blood b) rate of blood flow ii) if rate of blood flow approaches zero, amount of O2 available also approaches zero • Transport of Oxygen in dissolved state i) Normal arterial blood: 100 ml blood has dissolved 0.29 ml O2 [PO2 95 mmHg] ii) Venous blood: 100 ml blood has dissolved 0.12 ml O2 [PO2 40 mm Hg] iii) Thus, 0.17ml of O2 is transported by each 100 ml blood through the tissues per cycle in the dissolved state. B) Transport of Carbon dioxide in the blood • Normally 4 ml of CO2 is transported from the tissues to the lungs in each 100 ml blood • Gaseous CO2 (generally not bicarbonate) diffuses out of the cell • Chemical forms in which CO2 is transported: 1) Dissolved state [7%] i) arterial blood PCO2= 40 mmHg; 2.4 ml CO2 in 100 ml blood ii) venous blood PCO2=45 mm Hg; 2.7 ml CO2 in 100 ml blood iii) therefore, 1.3 ml is transported as dissolved CO2 by each 100 ml blood 2) Bicarbonate [70 %] i) reaction of CO2 with water in rbc—> carbonic acid ii) carbonic anhydrase catalyzes the reaction of CO2 & H2O 5000 X iii) carbonic acid —> H+ & HCO3- iv) H+ combines with Hb (Hb is a powerful acid-base buffer) v) HCO3- diffuse into plasma; Cl- diffuses into rbc [chloride shift] vi) administration of an carbonic anhydrase inhibitor —> reduced CO2 transport —> elevated tissue PCO2 3) Carbaminohemoglobin [23%] i) CO2 combines reversibly with Hb (and to a much lesser extent other plasma proteins) • Haldane effect i) is the effect of the oxygen-hemoglobin reaction on CO2 transport ii) binding of O2 with Hb tends to displace CO2 from the blood iii) Tissues: have increased CO2 uptake due to O2 removal from Hb iv) Lungs: have increased release of CO2 because of O2 pickup by Hb v) Due to increased acidity of Hb when combined with O2 vi) approximately doubles the amount of CO2 picked up in the tissues and released in the lungs • the formation of carbonic acid decreases the pH in venous blood [effect is attenuated by buffers]

References:

1.M.F. Perutz, 'Hemoglobin and Respiratory Structure', Scientific American,1978, 144-158'Allostery: Haemoglobin' (3 parts). 2.Bio inorganic and supramolecular chemistry.A.K. Bhagi and G.R. Chatwal. 3. Wilbur H. Campbell, 1995; wcampbel@mtu.edu 4. Haemoglobin - a molecular lung: 1. Peter E. Childs. 5. Bio Inorganic Chemistry, Lecture notes. Prof.(MS.) K.Day.2006.

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