1. Hemoglobin is an oxygen-carrying protein found in red blood cells. It is a tetramer composed of two alpha and two beta subunits that allows for cooperative oxygen binding. 2. Variants of hemoglobin can result in hemoglobinopathies like sickle cell anemia and thalassemia. Sickle cell anemia is caused by a single amino acid substitution that causes hemoglobin to polymerize under low oxygen conditions. 3. Thalassemia results from reduced or absent globin chain production leading to imbalanced globin synthesis and anemia. Haptoglobin is a protein that binds free hemoglobin in the bloodstream. It has alpha chain variants in humans that are the

1. CLINICAL ASPECTS OF BIOCHEMISTRY
PROTEINS AND DISEASE
HAEMOGLOBIN AND HAPTOGLOBIN
Haemoglobin - revision
Haemoglobin variants - haemoglobinopathies
Haemoglobin S
Thalassaemia
Haptoglobins

2. HAEMOGLOBIN - REVISION
Myoglobin (Mb) Oxygen binding/storage protein in muscle;
may also play a part in local oxygen transport. O2 binds to haem.
Maintenance of haem in Fe2+ form is necessary for O2 binding.
Mb is a monomeric protein of about 150 aa.
Haemoglobin A (HbA) O2 carrier in blood (red cells).
Tetramer: 22. Quaternary structure allows allostery - co-
operative binding of O2 modulated by pH (Bohr effect), CO2
binding, bisphosphoglycerate (BPG) binding. 3D structure of
each chain is similar to that of Mb.

3. HAEMOGLOBIN

4. TERTIARY STRUCTURE OF MYOGLOBIN AND
HAEMOGLOBIN  SUBUNIT

5. Different types of Hb in man:
HbA 22
HbA2 22 ~2% of adult Hb;  chain differs from  at ~ 10% of residues;
function (if any) unclear
HbF 22 late foetus and neonate; replaced by HbA 3-6 months after birth; 
chain differs from at ~of residues. In presence of BPG HbF has higher
affinity for O2 than HbA, allowing transfer of O2 to foetus (2 HbFs in man,  chains
differing at 1 aa)
HbGower Gower I 22 Embryonic.  similar to  (~20% differences)
Gower II 22 Embryonic.  similar to  (~40% differences)
So, at least 5 different Hbs (6 chains) in normal human. , , d,  chains can all
form tetramers,  can't.

6. DEVELOPMENTAL PATTERN OF HAEMOGLOBIN IN MAN
Based on Voet & Voet (1995)

7. EVOLUTIONARY TREE RELATING
HUMAN GLOBIN CHAINS
Mb







8. Hbs in lower organisms:
Mammals. Adult Hbs all similar to human HbA, but may be
variants unlike those seen in human. Developmental patterns
of Hbs differ considerably
Other vertebrates Most vertebrates have 22 type structure.
Variant types differ considerably. Lamprey (most primitive
fish) has only a single chain - more similar to Mb than
mammalian Hbs (no allostery)
Invertebrates, plants, bacteria. Hb-like proteins frequently
found, but not 22

9. HAEMOGLOBIN VARIANTS - HAEMOGLOBINOPATHIES
1. Exterior of molecule
e.g. Glu6Val [haemoglobin S (HbS)]
Glu B8Lys (harmless?)
2. Altered tertiary structure
Phe CD1Ser [Hb Hammersmith]
Gly B6Arg [Hb Riverdale-Bronx]
3. Altered 'active site’
His F8Tyr [Hb Iwate] (proximal His) )
His E7Tyr [Hb Boston] (distal His) )(cyanosis; methaemoglobinaemia)
4. Alterations at subunit interfaces
Asp G1His [Hb Yakima] )
Asn G4Thr [Hb Kansas] )
(polycythaemia or cyanosis)

10. SICKLE CELLANAEMIA - HbS - FIBRES
Based on Voet & Voet (1995)

11. HbS - AGGREGATION
Based on Voet & Voet (1995)

12. HAEMOGLOBIN S (HbS)
Possible therapies:
1. Disruption of intramolecular interactions (peptides?)
2. Use of agents to increase O2 binding affinity
3. Lower HbS concentration (increase erythrocyte
permeability)
4. Keep HbF switched on (hydroxyurea)
5. Vasodilators
6. Gene therapy

13.  and  THALASSAEMIAS
0 and 0 thalassaemias- corresponding globin chain missing completely
 and + thalassaemias - corresponding globin chain produced in reduced amount
 thalassaemia
1. Silent carrier state: 1 (of 4)  genes missing
. thalassaemia trait: 2  genes missing
3. Hb H disease: 3  genes missing
4. Hydrops fetalis: 4  genes missing [lethal]
[ Hb Barts: excess 4; HbH: excess 4 ]
Also  thalassaemia due to other causes. E.g. Hb Constant Spring:
Mutant stop codon and read-through of 31 aas, but mRNA degraded, so little
protein

14. HAEMOGLOBIN GENE CLUSTERS


 y y1 1 2
 G A y  
40
20 60 kbp
Chromosome 16
Chromosome 11

15. DELETIONS IN THE HAEMOGLOBIN  GENE CLUSTER
HPFH = hereditary persistent fetal haemoglobin
0 thalassaemia
Hb Lepore
GA 
thalassaemia
GA HPFH
G HPFH
Hb Kenya
 G A y  
40
20 60
kbp

16.  thalassaemia
Point mutations that can cause  thalassaemia:
1. Nonsense mutations
2. Frameshift
3. Point mutation in promoter
4. Point mutations that inactivate or generate splice sites
5. Point mutations of the AATAAA sequence

17. HAPTOGLOBINS
 &  chains; S-S linked; tend to form oligomers ()2 etc.
In human  chain is polymorphic:
I (83 residues): IF (Lys54) and IS (Glu54)
II (143 residues):
Lys Glu
54 71 113 143
Partial gene duplication
Gene frequencies: IF 0.16
IS 0.24
II 0.60

18. PROPOSED MECHANISM FOR PARTIAL GENE
DUPLICATION OF HAPTOGLOBIN


Hp1F
Hp1S
Hp2