The Functional Diversity of Proteins: The Example of Hemoglobin Flashcards
(45 cards)
Describe the overall secondary structure of myoglobin and hemoglobin chains.
Myoglobin is an all alpha helix protein with 153 amino acids. The oxygen-binding heme group (colored plane) is non-covalently bound to a pocket in the protein.
Hemoglobin and Myoglobin have very similar secondary structures they are both alpha helices. The differences, however, can be seen in their N- and C-terminus. The hemoglobin chain has more hydrophobic residues on its surface since it has to make contact with other subunits. The two of them are only 26% identical and 59% similar, however, their folding pattern is virtually identical.
Hemoglobin’s secondary structure has 11 and 22 structures that act as dimers. These two then form a tetramer.
Function of Myoglobin
Functions in oxygen storage in the skeletal muscle for contraction.
Function of Hemoglobin
Functions in oxygen transport from Lung to tissues.
Describe the quaternary structure of hemoglobin.
The quaternary structure of hemoglobin contains 4 heme-Fe2+ groups (1 per subunit) thus it can bind 4 O2 molecules. It is composed of four subunits, 2 of each kind (2 alpha, 2 beta). It is composed of 11 and 22 dimers that form a tetramer. Overall, there are 4 heme groups that allow 4 oxygen molecules to bind.
hemoglobinopathies
Hereditary disorders of hemoglobin
two main classes of hemoglobinopathies discussed:
- Structural Variants: These have an altered amino acid sequence. There are over 800 known. An example is sickle cell disease which results from a single amino acid mutation (Glu6Val)
- Thalassemias: These have a decreased abundance of one or more of the globin chain.
These are the most common single-gene diseases.
Indicate the mutation in sickle cell disease.
It is a point mutation in the beta chain and it is a point mutation of Glu6Val.
Describe the mechanism of red blood cell sickling and the symptoms that result.
Oxygenated the sickle cell hemoglobin (HbS) look similar to the normal hemoglobin. However, once they become deoxygenated they become “sickled”. This is a result of the formation of - aggregates. This aggregation occurs because the non-conservative mutation, or the replacement of a polar amino acid with a nonpolar one allows this new hydrophobic region to bind to a normally occurring hydrophobic patch on a normal hemoglobin molecule. This results in aggregation. Furthermore, the - interactions cause HbS to polymerize and to form long, insoluble rods of protein that result in deformation of the red blood cells, again causing aggregation. It is key to note that hemoglobin has to be deoxygenated for this to occur.
Indicate which conformational state the hemoglobin must be in for sickling to occur.
It must be in the deoxygenated form.
Describe the difference between sickle cell trait and sickle cell disease.
Those with the sickle cell trait are heterozygous for the autosomal recessive disorder (1 bad -chain gene and 1 good -chain gene). Thus, those with the trait only experience symptoms under extreme hypoxia. The heterozygotes are resistant to malaria and thus have a survival advantage. Those with the sickle cell disease (2 bad -chain genes). are those that are homozygous recessive, therefore having the disease.
Describe the geographical distribution of the sickle cell trait and the reason for this distribution.
There is a high percentage of individuals with the sickle cell trait in western Africa where malaria is prevalent.
Describe the two components of heme and the basic shape of heme.
Two components of the heme group are the 1) planar porphyrin structure and the 2) ferrous ion (Fe2+) in the middle with coordinated covalent bonds with the nitrogens of the 4 pyrrole rings. The overall structure is planar.
Point out the hydrophobic and hydrophilic edges of heme.
The hydrophobic regions are those in which are the hydrocarbons connected to the pyyrole rings. The hydrophilic regions are the propionic acid groups on the pyyrole ring.
Relate the oxidation state of the heme iron to oxygen binding.
The Fe2+ binds oxygen. If the Fe2+ becomes oxidized to Fe3+ then the oxygen can no longer bind to the iron. Hemoglobin that has the Fe3+ is called methemoglobin. The Fe2+ state must be present because this allows for one histidine to bind and one O2 molecule.
Describe the physicochemical properties of the heme binding pocket.
The heme binding pocket is between the E and F helices. The amino acids that make it up are hydrophobic.
Describe the consequences of the β-chain Phe42Ser mutation in Hb Hammersmith.
The hydrophobic Phe is mutated to a hydrophilic Ser. This smaller, hydrophilic amino acid allows the heme to slip out of its pocket due to the disruption of heme’s hydrophobic pocket. This mutated hemoglobin is also unstable and aggregates and precipitates. However, it does not form rods like HbS or sickle cell does.
What is the structure of oxygenated heme, indicating the 6 Fe2+ binding ligands?
Heme is bound to 4 pyyrole rings via the nitrogens. Then, one bond is with the proximal histidine or F8 histidine. The final bond is with Oxygen. The E7 distal histidine is not bound to the heme group, rather it forms a hydrogen bond with the bound oxygen molecule.
Describe the functions of the F8 proximal histidine and the E7 distal histidine.
The F8 proximal histidine forms a fifth coordinated covalent bond with the Fe2+. The E7 distal histidine does not make a bond to the iron, rather it h-bonds with the oxygen molecule. In the deoxygenated form, the iron is a little out of the plane, however, when it is oxygenated, the distal histidine’s h-bond to the oxygen helps pull the structure back into the plane. Furthermore, since the F8 proximal histidine is bound to the Fe and helix, it causes a conformational change in the protein.
Define the T-conformation and R-conformation with respect to oxygen binding.
T-conformation: Refers to the taut or tight form. It is in the deoxygenated form. It is “tight” so it can’t bind oxygen (not really true just a way to help remember that this is the deoxygenated form).
R-conformation: Refers to the “relaxed” form of hemoglobin. It is the oxygenated form of hemoglobin. The R form has 150 to 300-fold greater affinity for oxygen that T form.
Indicate which conformation has the highest affinity for oxygen.
The R-form has 150 to 300-fold higher oxygen affinity.
Explain positive cooperativity in oxygen binding to hemoglobin.
Positive cooperativity in regards to oxygen binding states that as oxygen binds to one of the 4 subunits of the hemoglobin, it will have a conformational change in that subunit (R-form) that will induce a conformational change on the other subunits to make them the R-form as well and have a higher affinity for oxygen. The opposite holds true for the dissociation as well. As one subunit loses and oxygen, it changes to the T-form, causing the other subunits to change to the T-form.
Indicate the trigger for the conformational change that occurs upon oxygen binding.
The trigger for the conformational change that occurs upon oxygen binding is the F8 proximal histidine, the oxygen molecule, and the E7 distal histidine. This is because in the deoxygenated form (T-form), the Fe2+ is a little bit out of the plane of the heme group. Therefore, when O2 binds to the Fe2+, the iron is brought more into the plane. Sine the F8 proximal histidine is bonded to the Fe2+, O2 causes this histidine to move as well. As part of the F helix, the F8 histidine then causes this F helix to change shape. This is at the FG corner.
Describe the consequence of the structural change in the FG corner when oxygen binds.
This conformational change in the FG corner that was described above affects the conformation of an adjacent subunit. The FG corner interacts with the C-helix of a neighboring subunit through non-covalent bonds interactions, such that a change in the FG corner changes the conformation of the C-helix toward the R-conformation. This interaction is mainly between the 12 and the 21 subunits.
Explain the consequence of the mutation β-chain Asp99Asn in Hb Kempsey.
When Asp or Aspartic acid which is negatively charges is mutated to Asn or Arginine which is basic and thus positively charged, we lose the negative charge that was previously present. This mutation at the FG corner locks the Hb in the high oxygen affinity state or the R-conformation. This causes the Hb to hold on to the Oxygen molecules.
