Haemoglobin Structure and Synthesis Flashcards
Main function of red cells
- carry O2 to tissues and return CO2 from tissues to lungs
What does each molecule of Hb A consist of?
- 4 polypeptide chains, alpha2 and beta2, each with its own haem group
What other 2 Hb’s does normal human blood contain?
- Hb F and Hb A2
What do these other 2 Hb’s contain?
- alpha chains with gamma and delta chains
Where does Haem synthesis occur?
- mitochondria
How does Haem synthesis commence?
- condensation of glycine and succinyl coenzyme A using key rate-limiting enzyme delta-ALA synthase
What is the coenzyme for this reaction?
- pyridoxal phosphate (vitamin B6)
What does protoporphyrin combine with to form haem?
- iron in the ferrous state
Haemoglobin molecular structure
- tetramer of 4 globin chains each w/ own haem group
What percentage of RBC volume and dry weight does Hb occupy?
- 33% RBC volume
- 90% dry weight
What 2 major components does a Hb molecule contain?
- globin chain
- prosthetic group Haem, comprising a tetrapyrolle ring structure w/ Fe(II) at its centre
Step 1 of Haem synthesis (ALA formation)
- succinyl CoA + glycine –> alpha amino beta ketoadipic acid
- CO2 is removed to form ALA
Step 2 of Haem synthesis (PBG/porphobilinogen formation)
- 2 ALAs
- ALA dehydratase is catalyst
- H20 removed to form PBG
Step 3 of Haem synthesis (polymerisation of PBG)
- 2 enzymes (PBG deaminase, Uroporphyrinogen III Consynthetase) work in unison
- 4 PBG form tetrapyrolle ring structure (Urogen III)
Step 4 of Haem synthesis (decarboxylation; acetyl to methyl)
- acetyl side chains converted to methyl groups through CO2 loss
- 4 CO2 liberated
- cytoplasmic structure: uroporphyrinogen III decarboxylase
Step 5 of Haem synthesis (conversion of proprionyl to vinyl)
- oxidative decarboxylation and dehydrogenation of 2 proprionyl groups, converts them to vinyl groups
- produces protoporphyrinogen IX
- enzyme: coproporphyrinogen oxidase
Step 6 of Haem synthesis (oxidation, removal of 6H+ atoms)
- mitochondrial enzyme; protoporphyrinogen oxidase
- responsible for removal of 6 H atoms
- produces protoporphyrin IX
Step 7 of Haem synthesis (insertion of Fe(II) into ring)
- insertion of ferrous, Fe2+ ion into centre of protoporphyrin IX
- inner mitochondrial enzyme; ferrochelatase or haem synthetase
Iron absorption and circulation
- Fe2+ readily absorbed by DMT1 by enterocytes
- Fe transported by transferrin in plasma to BM or liver
- transferrin; 76-80kD, carries 2 atoms of Fe3+
- ferroportin transports Fe out off cells
- carried more efficiently as Fe3+
- other carriers include albumin and lactoferrin
- Fe-Tr complex can only enter developing RBC by binding Transferrin receptor (TfR)
Hepcidin
- Movement of Fe into plasma by ferroportin regulated by it
- Produced by liver
- 25 amino acid peptide, HAMP gene, chromosome 19
- Controls export of Fe from; enterocytes, macrophages, Kupffer cells, hepatocytes, placental cells
- Raised hepcidin- anaemia of chronic disease
What is the Embden-Meyerhof pathway?
- glycolysis which converts glucose into pyruvate
- Free energy released forms ATP
- Oxygen free
- Due to lack of mitochondria within mature erythrocytes
What stage of red cell development does Hb synthesis occur?
- ~65% during the late normoblast stage
- 35% during the reticulocyte stage after loss of nucleus.
What does build up of Haem do?
- inhibits own synthesis and stimulates globin chain production
How much Hb does each normal red cell contain?
- 27-32pg
What is meant by ineffective erythropoiesis?
- red cell precursor fails to acquire sufficient Hb, so its destroyed before leaving the marrow
Primary structure of globin (globin genes)
- derived from 2 families of polypeptides – alpha & beta.
- Various Hb molecules produced contain 2 globins from each family.
- Synthesis of the globin chains is under control of globin genes, reside on the long arms of chromosomes 11 and 16
Globin genes
- between the active genes in these clusters are non-functional ‘pseudogenes’
- Two are duplicated; gamma and alpha.
- One gamma gene codes for alanine at position 136, the other coding for glycine at the same position.
- a chain gene is duplicated but both are identical and active.
Secondary structure of globin
- 75% of a and b globin chains are in form of a-helices
- All functional Hb molecules have this same helical content – 8 helices labelled A –> H.
- Remaining 25% of residues are in linear portions which connect the helices.
Tertiary structure of globin
- Haem group of each globin chain sits deep in a hydrophobic pocket or crevice between E and F helices.
- Proline residues within the linear portions of the globin give the chain necessary flexibility to take up its complex globular shape.
What does alteration in amino acid composition in Hb?
- may make Hb less stable and liable to precipitation.
- Precipitated Hb within a red cell renders it useless as an oxygen carrier and causes premature red cell destruction.
Glucose-6-phosphate dehydrogenase
- converts glucose-6 phosphate to 6-phosphoglucono-d-lactone and maintains NADPH
- G6PD deficiency can lead to lack of NADPH to maintain reduced glutathione to combat oxidant stress imposed on the cell
- Heinz Bodies; contain denatured Hb caused by oxidant damage
- Haemolytic anaemias
Structural contacts in a quaternary structure in globin
- Monomers held together by hydrophobic bonds between adjacent areas of the polymer.
- The monomers interact across various planes of contact.
- contacts mainly involve the B and H helices.
- These strong interactions stabilize the molecule and prevents dissociation of the a1b1 and a2b2 dimers into free monomers
Functional contacts in quaternary structure in globin
- The a1b2 and a2b1 contact planes are less extensive.
- Only 9 residues from either chain are involved.
- Involves contacts between C helix and the F-G corner
Functional contacts in O2 binding
- allow conformational changes of the molecule when the oxygenation state of Hb is changed.
- Deoxygenated Hb has the Fe(II) sitting slightly above the plane of the haem moiety.
- The haem is slightly concave due to interaction with F8
- Oxygenation leads to the movement of the Fe(II) into the plane of the porphyrin group.
- Fe(II) now sits within the plane of the porphyrin ring
- Simultaneous changes in the conformation of parts of the globins occurs.
What makes Hb an allosteric protein?
- Changes in shape of tetramer facilitate O2 uptake and release under physiological conditions.
- F helix and haem groups swing towards the centre of the molecule.
- Methaemoglobin (MetHb) contains Fe3+ in Haem. Cannot bind/exchange oxygen. MetHb Reductase pathway ensures MetHb is converted back into functional Hb
2,3-Biphosphoglycerate (2,3-BPG)
- binds to the b chains in the middle of the tetramer.
- combines with deoxyHb; reduces affinity of Hb for O2.
- Reversibly bound, altering the conformation of the Hb molecule, facilitating O2 release.
- O2 dissociation curve for Hb is shifted to the right-lower affinity.
- The greater the level of 2-3 BPG the more O2 is released from the Hb.
Factors that affect the release of O2 from Hb
- pH
- temperature
- PCO2
- PO2
- 2-3 BPG.
Foetal Hb
- Foetal Hb (HbF), has a higher affinity for O2 because it cannot bind 2-3 BPG strongly.
- Allows HbF to carry 20-30% more O2 than maternal HbA at a particular PO2
- HbF preferentially takes O2 from maternal Hb across the placental membranes.
