1. Oxygen Binding Proteins

2. Objectives: Hemoproteins carring O2 Myoglobin function and structure
Hemoglobin function and structure and forms
O2 binding to myoglobin and hemoglobin
O2 dissociation curve
Allosteric effectors

3. Why do we need oxygen carriers?
i. Cannot carry enough in blood to meet metabolic demand ii. Oxygen is very reactive – oxidizes iii. Oxygen cannot diffuse easily (we have thick skin)
Properties of a good oxygen carrier
i. Binds oxygen at a high [O2]
ii. Doesn’t oxidize cellular components
iii. Gives up oxygen on demand

4. Oxygen Binding Proteins are Hemoproteins
Oxygen binding proteins (myoglobin and hemoglobin) are hemoproteins
Hemoproteins are a group of specialized proteins that contain heme as a tightly bound prosthetic group.
Heme is a complex of protoporphyrin IX & ferrous iron (Fe2+) .
Fe2+ is held in the center of the heme molecule by bonds to the four nitrogens of the porphyrin ring.
It can form two additional bonds, one on each side of the planar porphyrin ring.
- one of these positions is coordinated to a histidine of globin
- the other position is available to bind oxygen

5. The heme group: Fe2+ porphyrin complex with bound O2
OXYGEN
PORPHYRIN IX
RING
FERROUS
ION
HISTIDINE
The heme group: Fe2+ porphyrin complex with bound O2

6. 1- Myoglobin Location: Structure:
heart & skeletal muscles giving them red color
Function:
a reservoir for oxygen for muscles. an oxygen carrier that increases the rate of transport of oxygen within the muscle cell.
Structure:
Myoglobin is composed of a single polypeptide chain that is structurally similar to the individual subunit polypeptide chain of hemoglobin.
The polypeptide chain is folded into 8 stretches labeled A to H

7. Myoglobin cont. Globin Protein (Polypeptide Chain) Heme group
The interior almost entirely of nonpolar
amino acids.
Polar amino acids are located almost
exclusively on the surface of the
molecule.
Heme group
is located in a pocket in the
molecule between helix E and
helix F which is lined by nonpolar
amino acids with exceptions
of two histidine amino acids.

8. Myoglobin cont.
The first is called the proximal histidine (F8), binds directly to the iron of heme.
The second is called the distal histidine (E7), does not directly interact with the heme group, but helps stabilize the binding of oxygen to the ferrous iron.
The protein, or globin, portion of myoglobin prevents the oxidation of iron of heme.

9. Myoglobin cont. Myoglobin as a diagnostic tool:
Myoglobin is used as a marker for acute myocardial infarction
Advantage:
It is elevated in blood of patients with myocardial infarction than other
markers as CK-MB or troponin. So, it can diagnose myocardial
infarction attack at an early stage.
Disadvantage:
myoglobin has a reduced specificity for diagnosing myocardial infarction.

10. 2- Adult Hemoglobin HbA1 Location: Function: Structure:
exclusively in red blood cells
Function:
main function is to transport
oxygen, H+ & CO2 and act as
RBCs buffer
Structure:
Hemoglobin is composed of four polypeptide chains - two α chains & two β chains – arranged into 2 dimers held together by noncovalent interactions.
Each subunit has 8 stretches of α-helical structure & a heme-binding pocket (as for myoglobin)
The oxygen-binding properties of hemoglobin are regulated by interaction with allosteric effectors

11. Quaternary structure of hemoglobin
The hemoglobin tetramer can be envisioned as being composed of two identical dimers, (αβ)1 & (αβ)2 (1 & 2 are numbers)
The two polypeptide chains within each dimer are held tightly together, primarily by hydrophobic interactions
In contrast, the two dimers are able to move with respect to each other, being held together primarily by polar bonds (ionic and hydrogen).
The weaker interactions between these mobile dimers result in the two dimers occupying different relative positions in deoxyhemoglobin as compared with oxyhemoglobin

12. Quaternary structure of hemoglobin cont.
TAUT
Structure T
RELAXED
Structure R
Structural changes resulting from oxygenation & deoxygenation of hemoglobin

13. Quaternary structure of hemoglobin cont.
T form:
The two αβ dimers interact through a network of ionic bonds that constrain the movement of the polypeptide chains.
The T form is the low-oxygen-affinity form of hemoglobin.
R form:
The binding of oxygen to hemoglobin causes the rupture of some of the ionic bonds between the αβ dimers.
This leads to a structure called the “R,” or relaxed form, in which the polypeptide chains have more freedom of movement
The R form is the high- oxygen-affinity form of hemoglobin.

14. Binding of oxygen to myoglobin & hemoglobin
Myoglobin can reversibly bind only one molecule of oxygen (O2), as it contains only one heme group.
Hemoglobin can bind four oxygen molecules (one at each of its four heme groups) cooperatively as 1st O2 bind at one heme increases the oxygen affinity of the remaining heme groups in the same hemoglobin molecule.
The degree of saturation (Y) of these oxygen-binding sites on all myoglobin or hemoglobin molecules can vary between zero (all sites are empty) and 100% (all sites are full

15. Oxygen dissociation curve
The oxygen dissociation curve is
a plot of saturation of binding sites with O2 in hemoglobin or myoglobin (Y) measured at
different partial pressures of oxygen (pO2)
Myoglobin has a higher oxygen affinity at any pO2 value than
hemoglobin as:
In myoglobin:
pO2 needed to achieve 50% saturation of
binding sites is ~ 1 mm Hg
In hemoglobin:
~ 26 mmHg

16. The ability of hemoglobin to reversibly bind oxygen is affected by:
Allosteric effectors
The ability of hemoglobin to reversibly bind oxygen is affected by:
pO2 pH
pCO2
2,3-bisphosphoglycerate (2,3 BPG) CO
These are collectively called allosteric (“other site”) effectors, because their interaction at one site on the hemoglobin molecule affects the binding of oxygen to heme groups at other locations on the molecule.
N.B.: The binding of oxygen to myoglobin is not influenced
by allosteric effectors.

17. 1- pO2 (Oxygen Concentration)
Loading & unloading of oxygen depend on pO2 (oxygen concentration
1- pO2 in alveoli of lungs (i.e. concentration of O2) is high
So, affinity of Hb to O2 is increased leading to saturation of hemoglobin with O2 (loading of oxygen)
2- pO2 in peripheral tissues is low
So, affinity of Hb to oxygen is decreased leading to release of oxygen to peripheral tissues (unloading of oxygen)
AT THE SAME TIME:
Myoglobin is designed to bind oxygen released from hemoglobin at low pO2
found in muscles (as myoglobin requires very low pO2 to be fully saturated
with oxygen)

18. 2- Bohr effect
Bohr effect is the change of oxygen binding in hemoglobin due to hydrogen ions (H+ or protons) and CO2
In peripheral tissues, H+ & CO2 are increased, which results to increased release of oxygen from hemoglobin.
1- H+ released from metabolism of peripheral tissues
2- CO2 resulting from cellular metabolism is converted by carbonic anhydrase to carbonic acid which is converted to bicarbonate & H+
In both cases, H+ increases the ionic bonds  favoring of T form of Hb   O2 release to the tissues
Hb O2 (OxyHb, R form) H ↔ HbH (deoxy Hb, T form) O2
So, increase in H+ shift equilibrium to right

19. 3- CO2
Most of CO2 produced during metabolism is hydrated and transported as bicarbonate ion
Some CO2 is carried as carbamate bound to the uncharged amino group
Hg-NH2+ CO Hb-NH-COO- + H+
This stabilizes the T –form and release O2 at tissuesbut in the lungs CO2 dissociates from Hb and released in breath.

20. 4- 2,3-bisphosphoglycerate (2,3 BPG)
Effect of 2,3-bisphosphoglycerate on oxygen affinity:
2,3- Bisphosphoglycerate (2,3-BPG) is an important regulator of the binding of oxygen to hemoglobin.
2,3-BPG is synthesized from an intermediate of the glycolysis.
Binding of 2,3-BPG to deoxyhemoglobin (T form):
2,3-BPG decreases the oxygen affinity of hemoglobin by binding to deoxyhemoglobin but not to oxyhemoglobin.
This preferential binding stabilizes the T conformation of deoxyhemoglobin.
Response of 2,3-BPG levels to chronic hypoxia or anemia:
The concentration of 2,3-BPG in the red blood cell increases in response to chronic
hypoxia (in certain lung diseases or high altitude) or chronic anemia
(oxygen available to Hb is low in these cases)
Elevated 2,3-BPG levels lower the oxygen affinity of hemoglobin, permitting greater unloading of oxygen in the capillaries of tissues.

23. 4- Carbon monoxide (CO)
Carbon monoxide (CO) binds tightly (but reversibly) to the hemoglobin iron, forming carboxyhemoglobin (carboxy Hb)
Dangers of CO binding to hemoglobin (in CO poisoning)
1- The affinity of hemoglobin for CO is 220 times greater than for oxygen (in availability of both, hemoglobin binds CO more)
2- When carbon monoxide binds to one or more of the four heme sites, hemoglobin shifts to the relaxed conformation (R-form), causing the remaining heme sites to bind oxygen with high affinity (tightly).
As a result, the affected hemoglobin is unable to release oxygen to the tissues leading to tissue hypoxia.
CO poisoning is treated with hyperbaric 100% oxygen therapy to facilitate the dissociation of CO from hemoglobin.

24. Factors favoring the T-form of Hb. are:
Deoxygenation
Low pH (  H+)
CO2
Lactic acid
2,3 bisphosphoglycerate
Factors favoring the R-form of Hb. are: O2 CO