lecture 7 haemoglobin

Definition: Hemoglobin (Hb) is a red globular protein pigment with a molecular weight of approximately 64,500. It is primarily found in red blood cells (RBCs).
Function: The main function of hemoglobin is to transport oxygen (O₂) from the lungs to the tissues.

Composition: Normal hemoglobin comprises globin, which is a tetramer consisting of two pairs of unlike globin chains (α and β chains). It contains four globular protein subunits, with each heme group featuring a protoporphyrin ring and a single iron ion (Fe²⁺).
Molecular Representation: The structure of hemoglobin can be represented with various chemical components including:
- Heme configuration
- Globin chains represented as α chain and β chain configurations.

Process: Heme and globin are synthesized separately within the developing red cell. The rates of heme and globin synthesis are equal to prevent excess of either.
Weight Contribution: Hemoglobin constitutes about 95% of the dry weight of red blood cells.
Synthesis Breakdown:
- 65% of hemoglobin is produced during the nucleated stages of RBC maturation.
- 35% occurs during the reticulocyte stage.
Heme Structure: Heme is classified as a porphyrin and consists of four pyrrole (C₄H₄NH) molecules linked via methane bridges, each pyrrole being coordinated to a ferrous ion.

Location of Synthesis: Heme synthesis occurs within the cytoplasm and mitochondria of red blood cells.
Key Steps:
- Initial Reaction: The first step, a rate-limiting step in heme synthesis, occurs with glycine and succinyl-CoA forming δ-aminolaevulinic acid (δ-ALA); it is energy-dependent and takes place in the mitochondria, catalyzed by a specific enzyme.
- Regulatory Mechanism: Free globin chains stimulate δ-ALA synthesis while free heme groups inhibit it, providing a control mechanism over heme synthesis.
- Cofactors: Cofactors like vitamin B6 derivatives (pyridoxal phosphate), ferrous, and copper ions are required for proper δ-ALA synthesis. The presence of free heme also inhibits the synthesis of the enzyme responsible for δ-ALA synthesis.
Intermediate Products:
- Two δ-ALA molecules combine asymmetrically to generate porphobilinogen (PBG) in a reaction facilitated by δ-ALA enzyme and glutathione.
- Subsequent reactions lead to condensed forms producing uroporphyrinogen III (UPG III) which then converts to coproporphyrinogen III (CPG III) through decarboxylation.
- CPG III subsequently enters mitochondria where it is converted to protoporphyrin IX (PPG IX) and ultimately to heme after iron insertion.

Polypeptide Chains: Various globins that combine with heme to form hemoglobin are single chain polypeptides whose synthesis is genetically controlled.
Gene Arrangement: There are eight functional globin genes:
- β-like Cluster: Contains β, γ, δ, and ε globin genes located on the short arm of chromosome 11.
- α-like Cluster: Contains α and ζ globin genes on the short arm of chromosome 16.
Globin Chain Variants: Different globin chains arise at various developmental stages (embryonic, fetal, adult). Key transitions include:
- Embryos synthesize ζ-chains and ε-chains.
- Over development, ζ-chain transitions to α-chain; ε-chain changes to γ-chain and then to β-chain.
Types of Hemoglobin Molecules:
- Embryonic Hemoglobins: Hb Gower 1 (ζ₂ε₂), Hb Portland (ζ₂γ₂), Hb Gower 2 (α₂ε₂), and fetal Hb (α₂γ₂).
- Normal Adult Hb: 96% Hb A (α₂β₂), 3% Hb A2 (α₂δ₂), and 1% fetal Hb (α₂γ₂).

Gas Transport: Hemoglobin's main roles include transporting O₂, CO₂, and H⁺ ions. Each hemoglobin molecule contains four heme groups.
Oxygen Loading Mechanism: When one hemoglobin heme binds O₂, it induces conformational changes enabling successive heme groups to bind O₂ more readily (termed the heme-heme interaction).
O₂ Affinity:
- The affinity of hemoglobin for O₂ is depicted through the O₂ dissociation curve, showing the relationship of O₂ partial pressure (pO₂) to hemoglobin saturation.
- The P50 value, the partial pressure of O₂ at which hemoglobin is 50% saturated, is normally around 26.6 mmHg.

Bohr Effect: Hemoglobin's affinity for O₂ is influenced by pH levels. A decrease in pH (becoming more acidic) leads to reduced O₂ affinity, facilitating O₂ release in tissues where acidity is heightened.
2,3-DPG Interaction: When hemoglobin releases O₂, β-chains separate, allowing 2,3-DPG (Diphosphoglycerate) to bind to deoxyhemoglobin, thus affecting O₂ affinity and transport.

Buffering Role: Hemoglobin participates as a significant blood buffer by binding H⁺ ions resulting from CO₂ dissolution in red blood cells, sustaining normal pH levels in the blood.
Carbon Dioxide Transport: The CO₂ from tissues enters red blood cells where it combines with water to form bicarbonate (HCO₃⁻) under the catalysis of carbonic anhydrase, which is crucial for CO₂ transport.

Overview: G-6-PD deficiency can result from factors like the consumption of fava beans or specific antibiotics, leading to hemolytic anemia due to the lack of NADPH in red cells, which lack mitochondria.
Role of NADPH: NADPH generated in the pentose phosphate pathway reduces disulfide forms of glutathione, essential for maintaining erythrocyte structural integrity and protecting against oxidative damage.
Oxidative Damage Mechanism: Accumulation of peroxides can induce disruption in red blood cell membranes, shortening their lifespan and promoting hemolysis.

Luebering-Rapaport Pathway: This pathway allows erythrocytes to synthesize and degrade 2,3-DPG, affecting O₂ affinity of hemoglobin through glycolysis.
Methaemoglobin Reductase Pathway: The metabolism of methaemoglobin is crucial, where the enzyme converts oxidized hemoglobin (MetHb) back to its ferrous form, which can bind O₂.

Function: While not directly producing ATP, this pathway generates NADPH (important for fatty acid and steroid synthesis) and ribose (essential for nucleic acids).
Usage: Rapidly dividing cells require these pentoses for nucleic acid synthesis, highlighting the importance of the pentose phosphate pathway in red cells and other high turnover tissues.