Hemoglobin

Hemoglobin and Iron Metabolism

Hemoglobin Normal Content Functions Structure Types Synthesis Fate Abnormal Hb Derivatives Iron Metabolism

1 Hemoglobin

Hemoglobin (Hb) is an iron-containing protein pigment that forms the coloring matter in red blood cells (RBCs).
  • It forms:
    • 95% of dry weight of RBC
    • 30–34% of wet weight of RBC
  • Hemoglobin is a chromoprotein.
  • Molecular weight → 68,000

Functions

  • Carries respiratory gases:
    • Oxygen
    • Carbon dioxide
  • Acts as a buffer.

2 Normal Hemoglobin Content

Average hemoglobin content in blood → 14–16 g/dL.

It varies according to:

  • Age
  • Sex
  • RBC count

According to Age

Age Hemoglobin Content
At birth Around 20 g/dL
After 3rd month 17–20 g/dL
After 1 year 10–14 g/dL
From puberty onwards 14–16 g/dL
At birth, in infants and growing children, hemoglobin content is high because of increased number of RBCs.

According to Sex

Adult males

15 g/dL

Adult females

14.5 g/dL

3 Functions of Hemoglobin

1. Transport of Respiratory Gases

Major function of hemoglobin → transport of respiratory gases.
  • Oxygen → lungs → tissues.
  • Carbon dioxide → tissues → lungs.
A. Transport of Oxygen
  • Oxygen binds with hemoglobin → oxyhemoglobin.
  • This process is called oxygenation.
  • Oxyhemoglobin is an unstable compound.
  • The combination is reversible.
  • Whenever oxygen is required, hemoglobin can release oxygen immediately.
B. Transport of Carbon Dioxide
  • Carbon dioxide binds with hemoglobin → carbaminohemoglobin.
  • It is also an unstable compound.
  • The combination is reversible → carbon dioxide can be released from the compound.
  • Affinity of hemoglobin for carbon dioxide → 20 times more than its affinity for oxygen.

2. Buffering Action

  • Hemoglobin acts as a buffer.
  • It plays an important role in acid-base balance.

4 Structure of Hemoglobin

  • Hemoglobin is a conjugated protein.
  • It is a heterotetrameric oxygen-binding protein.
  • Hemoglobin is made up of:
    • Globin → protein part
    • Heme → iron-containing pigment part.

Heme

  • Pigment part of heme → porphyrin.
  • Porphyrin is formed by four pyrrole rings (tetrapyrrole):
    • I
    • II
    • III
    • IV
  • Pyrrole rings are attached to one another by methane (CH₄) bridges.
  • Iron is attached to each pyrrole ring and globin molecule.
  • Iron in hemoglobin is present in ferrous (Fe²⁺) form.
  • It is present in an unstable or loose form.
  • In some abnormal conditions, iron is converted into ferric (Fe³⁺) state, which is a stable form.

Heme in Other Proteins

Myoglobin

Monomeric oxygen-binding protein present in muscles.

Neuroglobin

Monomeric oxygen-binding protein present in brain.

Globin

  • Globin contains 4 polypeptide chains:
    • 2 alpha (α) chains
    • 2 beta (β) chains
  • Each alpha polypeptide chain → 141 amino acid residues.
  • Each beta chain → 146 amino acid residues.

5 Types of Normal Hemoglobin

Hemoglobin is of 2 types:

  1. Adult hemoglobin (HbA)
  2. Fetal hemoglobin (HbF)
Replacement of fetal hemoglobin by adult hemoglobin:
  1. Starts immediately after birth.
  2. Is completed at about 10th–12th week after birth.

HbA and HbF differ structurally and functionally.

Structural Difference

Adult Hemoglobin — HbA
  • Globin contains:
    • 2 alpha polypeptide chains
    • 2 beta polypeptide chains
  • Formula → α₂β₂
Fetal Hemoglobin — HbF
  • Globin contains:
    • 2 alpha chains
    • 2 gamma chains instead of beta chains
  • Formula → α₂γ₂
  • Each gamma chain has 146 amino acid residues, like the beta chain.
Hemoglobin A₂ — HbA₂
  • HbA has a variant called Hemoglobin A₂ (HbA₂).
  • HbA₂ contains:
    • 2 alpha chains
    • 2 delta chains
  • Formula → α₂δ₂
  • Delta chain → 146 amino acid residues.
  • HbA₂ forms 2.5% (2–3%) of total hemoglobin.

Types of Normal Hemoglobin

Type Polypeptide chains in globin
HbA: Adult hemoglobin 2 α-chains + 2 β-chains (α₂β₂)
HbA₂: Adult hemoglobin 2 α-chains + 2 δ-chains (α₂δ₂)
HbF: Fetal hemoglobin 2 α-chains + 2 γ-chains (α₂γ₂)

Functional Difference

  • Fetal hemoglobin has more affinity for oxygen than adult hemoglobin.
  • Oxygen dissociation curve of fetal blood is shifted to the left.

6 Synthesis of Hemoglobin

  • Hemoglobin synthesis starts in the proerythroblastic stage of erythropoiesis.
  • Hemoglobin appears only in the intermediate normoblastic stage.
  • Hemoglobin synthesis continues until the reticulocyte stage.

Synthesis of Heme

Heme is synthesized from:

  • Succinyl-CoA
  • Glycine

Sequence of Events in Heme Synthesis

1
In the mitochondrion, 2 molecules of succinyl-CoA combine with 2 molecules of glycine and condense to form δ-aminolevulinic acid (ALA) by ALA synthase.
2
ALA is transported to the cytoplasm.
  • 2 molecules of ALA combine to form porphobilinogen in the presence of ALA dehydratase.
3
Porphobilinogen → uroporphobilinogen I by uroporphobilinogen I synthase.
4
Uroporphobilinogen I → uroporphobilinogen III by porphobilinogen III cosynthase.
5
From uroporphobilinogen III, a ring structure called coproporphyrinogen III is formed by uroporphobilinogen decarboxylase.
6
Coproporphyrinogen III is transported back to the mitochondrion, where it is oxidized to form protoporphyrinogen IX by coproporphyrinogen oxidase.
7
Protoporphyrinogen IX → protoporphyrin IX by protoporphyrinogen oxidase.
8
Protoporphyrin IX combines with iron to form heme in the presence of ferrochelatase.

Formation of Globin

  • Polypeptide chains of globin are produced in the ribosomes.
  • Four types of polypeptide chains:
    1. Alpha
    2. Beta
    3. Gamma
    4. Delta
  • Each globin molecule is formed by combination of 2 pairs of chains.
  • Adult hemoglobin → 2 alpha + 2 beta chains.
  • Fetal hemoglobin → 2 alpha + 2 gamma chains.

Configuration

  • Each polypeptide chain combines with one heme molecule.
  • After complete configuration, each hemoglobin molecule contains:
    • 4 polypeptide chains
    • 4 heme molecules.

7 Fate of Hemoglobin

After the lifespan of 120 days, RBC is destroyed by the reticuloendothelial system, particularly in the spleen.
  • Hemoglobin is released into plasma.
  • Hemoglobin is immediately degraded in reticuloendothelial cells and split into:
    1. Globin
    2. Iron
    3. Porphyrin
  • Globin → utilized for resynthesis of hemoglobin.
  • Iron → stored in the body.
  • Porphyrin → converted into biliverdin.
  • Most biliverdin → converted into bilirubin.
  • Bilirubin and biliverdin together are called bile pigments.

8 Normal Hemoglobin Derivatives

Hemoglobin derivative → altered hemoglobin formed when hemoglobin combines with a substance that is present in blood under normal physiological conditions.

The normal hemoglobin derivatives are:

  1. Oxyhemoglobin
  2. Reduced hemoglobin
  3. Carbaminohemoglobin
1. Oxyhemoglobin
  • Oxyhemoglobin (HbO₂) is formed when oxygen binds with hemoglobin in the lungs.
  • Oxygen binds with iron in the heme part of hemoglobin.
2. Reduced Hemoglobin
  • When oxyhemoglobin reaches the tissues:
  • Oxygen is released into the tissues.
  • Hydrogen combines with hemoglobin → reduced hemoglobin (HHb).
3. Carbaminohemoglobin
  • In the tissues, carbon dioxide binds with the globin part of hemoglobin.
  • It forms:
    • Carbaminohemoglobin
    • Carbohemoglobin.

9 Abnormal Hemoglobin

Abnormal types of hemoglobin or hemoglobin variants are pathological mutant forms of hemoglobin.
  • Variants are produced because of structural changes in polypeptide chains caused by mutations in genes of globin chains.
  • Most mutations do not produce serious problems.
  • Occasionally, some mutations result in disorders.

Abnormal hemoglobin is of 2 types:

  1. Abnormal hemoglobin in hemoglobinopathies
  2. Abnormal hemoglobin in thalassemia and related disorders.

1. Abnormal Hemoglobin in Hemoglobinopathies

Hemoglobinopathy → genetic disorder caused by abnormal polypeptide chains of hemoglobin.
  • It is due to mutation of the gene for any polypeptide chain.
  • This results in formation of a new variant of hemoglobin.

Abnormal hemoglobins include:

  • Hb S
  • Hb C
  • Hb E
  • Hb H
  • Hb M.

Types of Abnormal Hemoglobin

Abnormal hemoglobin Hemoglobinopathy caused Changes in hemoglobin structure
1. Hemoglobin S Sickle cell anemia Normal α-chains and abnormal β-chains; substitution of amino acid valine for glutamic acid at the sixth position of β-chains
2. Hemoglobin C Hemoglobin C disease Normal α-chains and abnormal β-chains; substitution of amino acid lysine for glutamic acid at the sixth position of β-chains
3. Hemoglobin E Hemoglobin E disease Normal α-chains and abnormal β-chains; substitution of lysine for glutamic acid at the 26th position of β-chains
4. Hemoglobin H Hemoglobin H disease (common type of α-thalassemia) α-chains are decreased, absent or abnormal
5. Hemoglobin M Hemoglobin M disease; blue baby syndrome in children characterized by cyanosis Hemoglobin is in the form of methemoglobin due to deficiency of cytochrome b5 reductase

2. Abnormal Hemoglobin in Thalassemia and Related Disorders

  • In thalassemia, different types of abnormal hemoglobin are present.
  • Polypeptide chains are:
    • Decreased
    • Absent
    • Abnormal
  • In α-thalassemia:
    • α-chains are decreased, absent or abnormal.
  • In β-thalassemia:
    • β-chains are decreased, absent or abnormal.
  • Hemoglobin H:
    • Causes hemoglobin H disease.
    • It is a common type of α-thalassemia.

Other Abnormal Types of Hemoglobin

  • Hemoglobin G
  • Hemoglobin I
  • Hemoglobin Bart’s
  • Hemoglobin Kenya
  • Hemoglobin Lepore
  • Hemoglobin Constant Spring.

10 Abnormal Hemoglobin Derivatives

Abnormal hemoglobin derivative → altered form of hemoglobin formed by combination of hemoglobin with substances other than oxygen and carbon dioxide.
  • Abnormal hemoglobin derivatives are formed by:
    • Carbon monoxide poisoning.
    • Combination of drugs such as:
      • Nitrites
      • Nitrates
      • Sulfonamides
      with hemoglobin.
  • High levels of abnormal hemoglobin derivatives in blood produce serious effects by preventing transport of oxygen.
  • This results in oxygen lack in tissues, which may be fatal.

Four types:

  1. Carboxyhemoglobin
  2. Methemoglobin
  3. Sulfhemoglobin
  4. Glycated hemoglobin.

A. Carboxyhemoglobin

  • Carboxyhemoglobin or carbonmonoxyhemoglobin is an abnormal hemoglobin derivative formed by combination of carbon monoxide with hemoglobin.
  • Carbon monoxide is a colorless and odorless gas.
  • Hemoglobin has 200 times more affinity for carbon monoxide than oxygen.
  • Therefore, carbon monoxide hinders oxygen transport, resulting in tissue hypoxia.

Sources of Carbon Monoxide

  1. Charcoal burning.
  2. Coal mines and deep wells.
  3. Underground drainage system.
  4. Exhaust of gasoline engines.
  5. Gases from guns and other weapons.
  6. Heating system with poor or improper ventilation.
  7. Smoke from fire.
  8. Tobacco smoking.

Signs and Symptoms of Carbon Monoxide Poisoning

1. While breathing air with less than 1% carbon monoxide
  • Hemoglobin saturation → 15–20%
  • Mild symptoms such as:
    • Headache
    • Nausea
2. While breathing air with more than 1% carbon monoxide
  • Hemoglobin saturation → 30–40%
  • Severe symptoms such as:
    • Convulsions
    • Cardiorespiratory arrest
    • Unconsciousness
    • Coma
When hemoglobin saturation increases above 50%:

Death occurs.

B. Methemoglobin

  • Methemoglobin is an abnormal hemoglobin derivative formed when the iron molecule of hemoglobin is oxidized from the normal ferrous state → ferric state.
  • Methemoglobin is also called ferrihemoglobin.
  • Normal methemoglobin level → less than 3% of total hemoglobin.

Sources of Methemoglobin

  • Contaminated well waters with:
    • Nitrates
    • Nitrites
  • Matchstick explosives.
  • Naphthalene balls.
  • Nitrous oxide.
  • Irritant gases.

Methemoglobinemia

  • Methemoglobinemia → disorder characterized by a high level of methemoglobin in blood.
  • It leads to tissue hypoxia.
  • It causes:
    • Cyanosis
    • Other symptoms.

C. Sulfhemoglobin

  • Sulfhemoglobin is an abnormal hemoglobin derivative formed by combination of hemoglobin with hydrogen sulfide.
  • It is caused by drugs such as sulfonamides.
  • Normal sulfhemoglobin level → less than 1% of total hemoglobin.

D. Glycated Hemoglobin

  • Glycated hemoglobin (HbA1c) or glycosylated hemoglobin is formed when sugar gets attached with hemoglobin.
  • HbA1c test measures the average blood sugar level 2–3 months before the test.
  • Normal glycated hemoglobin level → less than 5.7%.

Formation of Abnormal Hemoglobin Derivatives

Derivative Formation
1. Carboxyhemoglobin By combination of hemoglobin with carbon monoxide
2. Methemoglobin By oxidation of hemoglobin from ferrous state to ferric state
3. Sulfhemoglobin By combination of hemoglobin with hydrogen sulfide
4. Glycated hemoglobin By combination of hemoglobin with sugar

11 Iron Metabolism

Importance of Iron

Iron is an essential mineral and an important component of proteins involved in oxygen transport.
  • Iron is important for formation of:
    • Hemoglobin
    • Myoglobin
  • Iron is also necessary for formation of:
    • Cytochrome
    • Cytochrome oxidase
    • Peroxidase
    • Catalase.

Normal Values and Distribution of Iron in the Body

  • Total quantity of iron in the body → about 4 g.
  • Each gram of hemoglobin contains 3.34 mg of iron.

Approximate Distribution of Iron in the Body

Location Iron Distribution
1. In blood 50–150 μg/dL
2. In hemoglobin 65–68%
3. In muscle as myoglobin 4%
4. As intracellular oxidative heme compound 1%
5. In plasma as transferrin 0.1%
6. Stored in reticuloendothelial system 25–30%

Dietary Iron

Dietary iron is available in 2 forms:

1. Heme Iron

Heme iron is present in:

  • Fish
  • Meat
  • Chicken

Heme iron is absorbed easily from the intestine.

2. Nonheme Iron

Nonheme iron is available in:

  • Vegetables
  • Grains
  • Cereals

Nonheme iron is not absorbed as easily as heme iron.

Cereals, flours and products of grains enriched or fortified with iron become good dietary sources of nonheme iron, particularly for:
  • Children
  • Women.

Absorption of Iron

  • Iron is absorbed mainly from the small intestine.
  • It is absorbed through intestinal cells (enterocytes) by pinocytosis and transported into the blood.
  • Bile is essential for absorption of iron.
  • Iron is present mostly in ferric (Fe³⁺) form.
  • It is converted into ferrous (Fe²⁺) form, which is absorbed into the blood.

Transport of Iron

  • Immediately after absorption into blood:
  • Iron combines with a β-globulin called apotransferrin.
  • Apotransferrin is secreted by the liver.
  • This results in formation of transferrin.
  • Iron is transported in blood in the form of transferrin.
  • Iron combines loosely with globin and can be released easily at any region of the body.

Storage of Iron

  • Iron is stored in large quantities in:
    • Reticuloendothelial cells
    • Hepatocytes in liver
  • It is also stored in other cells, but in small quantities.
In the cytoplasm of the cell:
  • Large amount of iron → stored as ferritin
  • Small quantity of iron → stored as hemosiderin.

Daily Loss of Iron

In Males

About 1 mg of iron is excreted every day through feces.

In Females

In adult females, iron loss is high because of menstruation.

12 Iron Content Related to Hemoglobin

Each gram of hemoglobin contains 3.34 mg of iron.

Normally, 100 mL of blood contains:

  • 15 g hemoglobin
  • About 50 mg iron (3.34 × 15)
Blood loss:

Therefore, if 100 mL of blood is lost:

About 50 mg of iron is lost.

During Menstrual Cycle

  • In females, during every menstrual cycle:
    • About 50 mL of blood is lost.
    • About 25 mg of iron is lost.
  • Therefore, iron content is always less in females than in males.

During Hemorrhage and Blood Donation

  • Iron is also lost during:
    • Hemorrhage
    • Blood donation
  • If 450 mL of blood is donated:
    • About 225 mg of iron is lost.

13 Regulation of Total Iron in the Body

Absorption and excretion of iron are maintained almost equally under normal physiological conditions.
  • When iron storage is saturated in the body:
    • Further absorption of iron from the gastrointestinal tract is automatically reduced.
  • This occurs by a feedback mechanism.

14 Applied Physiology: Iron Deficiency Anemia

  • Deficiency of iron causes a decrease in hemoglobin synthesis.
  • This results in iron deficiency anemia.
Hemoglobin and Iron Metabolism