Heamoglobin

Question 1

What type of metabolism does haemoglobin play a role in?

Answer 1

Aerobic respiration - involved primarily in transfer of Oxygen.

Question 2

What determines the binding of oxygen to Haemoglobin?

Answer 2

The haem groups. it’s not part of the polypeptide chain however, it’s tightly bound to the protein.

Question 3

Describe the structure of Haemoglobin.

Answer 3

Haemoglobin is a quartenary structure with 4 polypeptide chains. It can bind 4 oxygens. These subunits are held together by non-covalent interactions.

Question 4

What non covalent interactions are involved in Haemoglobin?

Answer 4

1- VDW interactions.
2- Electrostatic forces.
3- Hydrogen bonds.

Question 5

What is the function of Haemoglobin?

Answer 5

1- To transport Oxygen to tissues.
2-To transport coz and protons away from tissues.

Question 6

Discuss the schematics of oxygen binding site.

Answer 6

The site at which oxygen binds in both Hb and myoglobin is the haem which is found in the hydrophobic pocket of the protein. At the center of this is an iron atom held in place by 4 nitrogens.

Question 7

What holds the haem groups in the correct position within the hb chain?

Answer 7

The proximal histidine (f8) directly bonds to the iron in the haem and pulls it out of the frame. To prevent complete dislocation of the iron, and oxygen molecule binds also to the iron and pulls it back to the plane. This oxygen molecule is also bonded to the distal histidine (E7) which also contributes to the position of the haem.

Question 8

What is myoglobin?

Answer 8

Oxygen reservoir within the heart and skeletal muscles cells.
Consists of 8 helices- A to H.
Non helical region= A to B and B to C.

Question 9

Which of the 2 has a higher affinity for oxygen: myoglobin or haemoglobin?

Answer 9

Myoglobin- it doesn’t have a quaternary structure (monomer) therefore binding oxygen more tightly that hb. This difference in binding energy results in a hyperbolic shape in the oxygen dissociation curve.

Question 10

What are the variant forms of hb and how are they expressed?

Answer 10

HbA- adult
HbA2- minor adult form
HbF-foetal form
Hb Gower1- embryonic
Hb Gower2- embryonic
Hb Portland- embryonic

Question 11

What is sickle haemoglobin?

Answer 11

Arises from the single substitution of the amino acid glutamate to valine in hb. This causes hb molecules to distort and form a sickle shape.

Question 12

What are the effects of sickle cell anaemia?

Answer 12

1- Distorted shape leads to stacking of HbS into long fibres at low oxygen concentrations.
2- RBC looses its flexibility and affinity for oxygen drops.
3- Results in blockage of capillaries which can result in anoxia in tissues (leads to cell death).
4- Typically results in infection, dehydration, cold and hypoxia.

Question 13

Describe the clinical consequences of sickle cell anaemia.

Answer 13

1-Bone pain resulting from blockages.
2-Chronic anaemia- destruction of erythrocytes.
3-Cerebrovascular accidents.
4-Organ damage (kidneys, heart and lungs).
5-Acute crisis- very painful.

Question 14

What are the methods of diagnosing sickle cell anaemia and what are probable treatments?

Answer 14

Diagnosing: protein or genetic analysis.
Treatment:
1- most symptom management (hydration and analgesics).
2- Aggressive abx.
3- Blood transfusion (grey area as it can lead to iron overload).
4- Hydroxyurea.

Question 15

Discuss hydroxyurea as treatment option for sickle cell anaemia.

Answer 15

Anti cancer drug primarily. Changes gene expression to increase levels of foetal hb which is not normally expressed in adults. Foetal hb has a higher affinity for oxygen and it’s levels are ^ by approx 40%.

Question 16

Discuss the binding of oxygen to hb.

Answer 16

Oxygen and hb are known to have a cooperative binding relationship, meaning that for every oxygen that is bound to hb, it’s affinity for O2 increases. This results in a sigmoid curve on the dissociation graph.
*The affinity of the 4th bound O2 is 300 times greater than the 1st bound O2.

Question 17

How does hb dissociate?

Answer 17

At higher partial pressure (more than Mb). This allows for delivery of O2 from hb to mb.

Question 18

Describe the 2 structural changes to hb due to oxygenation.

Answer 18

Tense form: deoxy hb. Has a low affinity for O2. Structure is constrained by 8 salt bridges.
Relaxed form: Oxy hb. Higher affinity for O2.

Question 19

How is hb an allosteric protein?

Answer 19

Due to its cooperative relationship: the binding of O2 to one of its subunits is affected by its interactions with other subunits.

Question 20

What are the allosteric effects of hb?

Answer 20

Haem- haem interactions.
The Bohr effects.
2,3 Bisphosphoglycerate.

Question 21

What is the Bohr effect.

Answer 21

At low ph (or ^CO2), O2 is released more easily. This shifts the dissociation curve to the right (^p50). This maximises efficient handling of O2.

Question 22

What causes the Bohr effect?

Answer 22

^pCO2 leads to decrease in pH. Follow steps below;
1- CO2 binds to H2O and creates carbonic acid (H2CO3) in presence of carbonic anhydrase.
2- H2CO3 deprotonates and creates bicarbonate. This free H+ lowers pH.
*^H+ typically favours deoxy form.

Question 23

How are the H+ utilised?

Answer 23

They protonate some histidines and and some N-term alpha amino groups allowing them to form salt bridges which stabilises the deoxy Hb (T) form. As a result, ^H+ favours T form!

Question 24

In what form is CO2 transported?

Answer 24

70% as bicarbonate and 30% as carbamate in erythrocytes.
Carbamate form salt bridges and favour T form.

Question 25

What is 2,3 biphosphoglycerate?

Answer 25

It’s a byproduct of glycolysis and is present in erythrocytes in equimolar concentrations to hb. It binds to deoxy hb only thereby decreasing its affinity for for O2. This stabilises the taut T conformation (encourages it to deposit last big of O2).
*Converted from 1,3 BPG by biphosphogylcerate mutase.

Question 26

Describe Oxygenation in the presence of high levels of 2,3 BPG. When do the levels of 2,3 BPG ^?

Answer 26

2,3 BPG has 4 neg charges that form salt bridges with pos charged residues in the beta subunits within the central cavity of deoxy hb. Salt bridges must be broken during oxygenation (narrowing of cavity and ejection of 2,3 BPG).
^BPG occurs under hypoxic conditions e.g high altitudes, anaemia or emphysema.

Question 27

What happens to delivery of O2 in tissues if levels of 2,3 BPG is increased?

Answer 27

Delivery of O2 goes up, more O2 dissociates at higher PO2.

Question 28

What is anaemia?

Answer 28

A decrease in hb concentration below the reference range for age and sex.
Female: 11.5- 16 g/dl
Male: 13.5- 17.5 g/dl

Question 29

Name 2 inherited haemolytic anaemias.

Answer 29

Glucose 6 phosphate dehydrogenase deficiency (G6PD)- only some mutations cause symptoms.
Sickle cell anaemia

Question 30

How is the G6PD gene inherited and what does it cause?

Answer 30

Through the x linked chromosome.
It impairs the ability of an erythrocytes to form NADPH, resulting in haemolysis.
It also prevents the regeneration of glutathione and increases the Oxidative damage.

Question 31

What is NADPH and what is its role?

Answer 31

*NADPH- nicotinamide adenine dinucleotide phosphate. It’s a coenzyme that acts as a donor or accept of electrons. It minimises the effect of reactive Oxygen species that are produced in aerobic metabolism.
It also helps reduce glutathione (G-SH) as oxidised form (G-S-S-G) has no protective effect from oxidative damage.

Question 32

What are the factors that increase oxidative stress?

Answer 32

Antibiotics
Antimalarials
Antipyretics
Favism- fave beans
Infection and subsequent inflammatory response- generation of free radicals.
Neonatal jaundice- impaired catabolism of haem or ^ production of bilirubin.