2. • Hemoglobin derivatives are formed by the
combination of different ligands with the
heme part, or change in the oxidation state
of iron.
3. Carboxy-Hemoglobin (CO-Hb)
• Hemoglobin binds with carbon monoxide
(CO) to form carboxy-Hb.
• The affinity of CO to Hb is 200 times more than
that of oxygen.
• It is then unsuitable for oxygen transport.
4. • When one molecule of CO binds to one
monomer of the hemoglobin molecule, it
increases the affinity of others to O2; so that
the O2 bound to these monomers are not
released.
• This would further decrease the availability
of oxygen to the tissues.
5. Carbon Monoxide Poisoning
• CO is a colorless, odorless, tasteless gas
generated by incomplete combustion.
• CO poisoning is a major occupational hazard
for workers in mines.
• Breathing the automobile exhaust in closed
space is the commonest cause for CO
poisoning
6. • The carboxy-Hb level in normal people is
0.16%.
• An average smoker has an additional 4% of
CO-Hb.
• One cigarette liberates 10–20 ml carbon
monoxide into the lungs.
7. Clinical Manifestations
• Clinical symptoms manifest when carboxy-Hb
levels exceed 20%.
• Breathlessness, headache, nausea, vomiting,
& chest pain.
• At 40-60% saturation, death can result.
• Administration of O2 is the treatment.
8. Methemoglobin (Met-Hb)
• When the ferrous (Fe2+ ) iron is oxidized to
ferric (Fe3+) state, met-Hb is formed.
• Small quantities of met-Hb formed in the RBCs
are readily reduced back to the ferrous state
by met-Hb reductase enzyme systems.
• About 75% of the reducing activity is due to
enzyme system using NADH & cytochrome b5
9. Methemoglobinemias
• Normal blood has only less than 1% of
methemoglobin.
• It has markedly decreased capacity for
oxygen binding and transport.
• An increase in methemoglobin in blood,
(methemoglobinemia) is manifested as
cyanosis.
• Causes may be congenital or acquired.
10. Congenital Methemoglobinemia
• Presence of Hb variants like HbM can cause
congenital methemoglobinemia.
• Cytochrome b5 reductase deficiency is characterized
by cyanosis from birth.
• 10-15% of hemoglobin may exist as methemoglobin.
• Oral administration of methylene blue, 100-300
mg/day or ascorbic acid 200-500 mg/day decreases
met-Hb level to 5-10% and reverses the cyanosis.
11. Acquired or Toxic Methemoglobinemia
• Met-hemoglobinemia may develop by intake
of water containing nitrates or due to
absorption of aniline dyes.
• Drugs which produce met-hemoglobinemia -
acetaminophen, phenacetin, sulphanilamide,
amyl nitrite, & sodium nitroprusside.
12. Sulf-hemoglobinemia
• When hydrogen sulfide acts on oxy-Hb, sulf-hemoglobin
is produced.
• It occur in people taking drugs like
sulphonamides, phenacetin, acetanilide,
dapsone, etc.
• It cannot be converted back to oxy-hemoglobin.
13. Hemoglobinopathies
• Abnormal hemoglobins are the resultant of
mutations in the genes that code for α or β
chains of globin
• As many as 400 mutant hemoglobins are
known.
• About 95% of them are due to alteration in
single amino acid of globin
14. Types of abnormal Hb
• Two types:
• If the mutation affects structural gene, it
results in replacement of a single amino acid
in Hb by some other amino acid resulting into
abnormal Hb.
• E.g: Hb-S, Hb-M, Hb-C, Hb-D & others.
15. • If the mutation affects the regulator gene,
which affects the rate of synthesis of
peptide chains, the amino acid sequence
remains unaffected.
• E.g: Thalassaemias
16. Globin synthesis
• The globin genes are organised into two gene
families or clusters
• α-Gene family:
• There are 2 genes coding for α-globin chain
present on each one of chromosome 16.
• The ζ (zeta)-gene, other member of a-gene
cluster is also found on chromosome 16 & is
active during the embryonic development
17. • β-Gene family:
• The synthesis of β-globin occurs from a single
gene located on each one of chromosome 11.
• This chromosome also contains four other
genes.
• One ε-gene expressed in the early stages of
embryonic development.
18. • Two γ-genes (Gγ & Aγ) synthesize γ-globin
chains of fetal hemoglobin (HbF).
• One δ-gene producing δ-globin chain found in
adults to a minor extent (HbA2).
19. Sickle-cell anemia (HbS)
• Sickle-cell anemia (HbS) is the most common
form of abnormal hemoglobins.
• Erythrocytes of these patients adopt a sickle
shape (crescent like) at low oxygen
concentration
• It primarily occurs in the black population.
20. Molecular basis of HbS
• The glutamic acid in the 6th position of β chain
of HbA is changed to valine in HbS.
• This single amino acid substitution leads to
polymerization of hemoglobin molecules
inside RBCs.
• This causes a distortion of cell into sickle
shape
22. • The substitution of hydrophilic glutamic acid
by hydrophobic valine causes a localized
stickiness on the surface of the molecule
• The deoxygenated HbS may be depicted with
a protrusion on one side and a cavity on the
other side, so that many molecules can
adhere and polymerize
23. • The sickled cells form small plugs in
capillaries.
• Occlusion of major vessels can lead to
infarction in organs like spleen.
• Death usually occurs in the second decade of
life.
24. Homozygous and heterozygous HbS
• Sickle cell anemia is said to be homozygous, if
caused by inheritance of two mutant genes
(one from each parent) that code for β-chains.
• In case of heterozygous HbS, only one gene (of
β-chain) is affected while the other is normal
25. • The erythrocytes of heterozygotes contain
both HbS & HbA & the disease is referred to as
sickle cell trait.
• The individuals of sickle-cell trait lead a normal
life, & do not usually show clinical symptoms.
26. Abnormalities associated with HbS
• Life-long hemolytic anemia:
• The sickled erythrocytes are fragile & their
continuous breakdown leads to life-long
anemia.
• Tissue damage and pain:
• The sickled cells block the capillaries resulting
in poor blood supply to tissues.
• This leads to extensive damage & inflammation
of certain tissues causing pain.
27. • Increased susceptibility to infection :
• Hemolysis & tissue damage are accompanied
by increased susceptibility to infection &
diseases.
• Prematured eath:
• Homozygous individuals of sickle-cell anemia
die before they reach adulthood (< 20 years)
28. Mechanism of sickling in sickle-cell anemia
• Glutamate is a polar amino acid & it is
replaced by a non-polar valine in sickle-cell
hemoglobin.
• This causes a marked decrease in the solubility
of HbS in deoxygenated form
• Solubility of oxygenated HbS is unaffected
29. Sticky patches & formation of
deoxyhemoglobin fibres
• The substitution of valine for glutamate
results in a sticky patch on the outer surface
of β-chains.
• It is present on oxy- & deoxyhemoglobin S
but absent on HbA.
• There is a site or receptor complementary to
sticky patch on deoxyHbS.
30. • The sticky patch of one deoxyHbS binds with
the receptor of another deoxyHbS & this
process continuous resulting in the formation
of long aggregate molecules of deoxyHbS
• The polymerization of deoxy-HbS molecules
leads to long fibrous precipitates.
31. • These stiff fibres distort the erythrocytes into
a sickle or crescent shape
• The sickled erythrocytes are highly
vulnerable to lysis.
• ln case of oxyHbS, the complementary
receptor is masked, although the sticky patch
is present.
32. HbS gives protection against malaria
• HbS affords protection against Plasmodium
falciparum infection
• Hence the abnormal gene was found to offer
a biologic advantage.
34. Diagnosis of sickle cell anemia
• Sickling test:
• A simple microscopic examination of blood
smear prepared by adding reducing agents
such as sodium dithionite.
• Sickled erythrocytes can be detected under
the microscope
35. Electrophoresis
• Electrophoresis at alkaline pH shows a slower
moving band than HbA.
• At pH 8.6, carboxyl group of glutamic acid is
negatively charged.
• Lack of this charge on HbS makes it less negatively
charged, & decreases the electrophoretic mobility
• At acidic pH, HbS moves faster than HbA.
• In sickle cell trait, both the bands of HbA and HbS can
be noticed
37. Management of sickle cell disease
• Administration of sodium cyanate inhibits
sickling of erythrocytes
• Cyanate increases the affinity of O2 to HbS &
lowers the formation of deoxyHbS
• It causes certain side effects like peripheral
nerve damage
• In severe anemia, repeated blood transfusion
is required.
• It result in iron overload & cirrhosis of liver
38. Hemoglobin C disease
• Cooley's hemoglobinemia (HbC) is characterized by
substitution of glutamate by lysine in the sixth position
of β-chain.
• Due to the presence of lysine, HbC moves more slowly
on electrophoresis compared to HbA and HbS.
• HbC disease occurs only in blacks.
• Both homozygous & heterozygous individuals of HbC
disease are known.
• It is characterized by mild hemolytic anemia.
• No specific therapy is recommended.
39. Hemoglobin D
• Caused by the substitution of glutamine in
place of glutamate in the 121st position of β-
chain.
• Several variants of HbD are identified from
different places indicated by the suffix.
• For instance, HbD (Punjab)
• HbD, on electrophoresis moves along with
HbS.
40. Hemoglobin E
• Most common abnormal hemoglobin after HbS.
• lt is estimated that about 10% of the population in
South-East Asia (Bangladesh, Thailand, Myanmar)
suffer from HbE disease.
• In India, it is prevalent in West Bengal.
• HbE is characterized by replacement of glutamate by
lysine at 26th position of β-chain.
• The individuals of HbE (either homozygous or
heterozygous) have no clinical manifestations
41. Thalassemias
• Thalassemias are a group of hereditary
hemolytic disorders characterized by
impairment/imbalance in the synthesis of globin
chains of Hb
• Thalassemias (Greek: thalassa-sea) mostly
occur in the regions surrounding the
Mediterranean sea, hence the name.
• Also prevalent in Central Africa, India.
42. Molecular basis of thalassemias
• Hemoglobin contains 2α & 2β globin chains.
• The synthesis of individual chains is so
coordinated that each α-chain has a β-chain
partner & they combine to finally give
hemoglobin (α2β2).
• Thalassemias are characterized by a defect in
the production of α-or β-globin chain
43. • Thalassemias occur due to a variety of
molecular defects
• Gene deletion or substitution,
• Underproduction or instability of mRNA,
• Defect in the initiation of chain synthesis,
• Premature chain termination.
44. α-Thalassemiasas
• α-Thalassemias are caused by a decreased
synthesis or total absence of α-globin chain of
Hb.
• There are four copies of α-globin gene, two on
each one of the chromosome 16.
• Four types of α-thalassemias occur which
depend on the number of missing α-globin
genes
45. Salient features of different α -thalassemias
• Silent carrier state is due to loss of one of the
four α -globin genes with no physical
manifestations.
• α -Thalassemia trait caused by loss of two genes
(both from the same gene pair or one from each
gene pair).
• Minor anemia is observed
46. • Hemoglobin H disease, due to missing of three
genes, is associated with moderate anemia
• Hydrops fetalis is the most severe form of α-
thalassemias due to lack of all the four genes.
• The fetus usually survives until birth & then dies.
47. β-thalassemias
• Decreased synthesis or total lack of the
formation of β-globin chain causes β-
thalassemias.
• The production of α-globin chain continues to
be normal, leading to the formation of a globin
tetramer (α4) that precipitate.
• This causes premature death of erythrocytes.
• There are mainly two types of β-thalassemias
48. β-Thalassemia minor
• This is an heterozygous state with a defect in
only one of the two β-globin gene pairs on
chromosome 11.
• Also known as β -thalassemia trait, is usually
asymptomatic, since the individuals can make
some amount of β-globin from the affected
gene
49. β-Thalassemia major
• This is a homozygous state with a defect in
both the genes responsible for β-globin
synthesis.
• The infants born with β-thalassemia major
are healthy at birth since β-globin is not
synthesized during the fetal development
50. • They become severely anemic and die within
1-2 years.
• Frequent blood transfusion is required for
these children.
• This is associated with iron overload which in
turn may lead to death within 15-20 years
51. References
• Text book of Biochemistry – U Satyanarayana
• Text book of Biochemistry – DM Vasudevan
• Text book of Biochemistry – MN Chatterjea
