2. Hemoglobin and Red Blood Cells
Hemoglobin quaternary structure,
Contains 2a and 2b subunits
At max. capacity, the average human’s blood cells can hold over a liter of oxygen

4. Heme in Hemoglobin (Hb) a Myoglobin (Mb)
Myglobin is a monomeric protein (only 1 protein) and contains 1 heme.
Heme
Hemoglobin is a tetrameric protein (4 proteins) and contains 4 hemes.

5. Deoxyheme and Oxyheme Fe2+ + O2 Fe2+ (5 ligands bound)
= True color in O2 poor blood
= True color in O2 rich blood
For many years, scientists thought the veins where blue do to oxygen poor blood. We no longer believe this is the case, but that there is an optical effect.
Many types of Iron Oxides are red. Consequently, it is the iron in blood which cause it to be red.

6. MRI + Hemoglobin has allowed us to view the Human Brain in Action
The affect to the left has been triggered by a man sensing a smell. The regions of the brain consuming O2 are primarily of the olfactory cortex which can be visualized do to differences in the MRI between oxy and deoxy hemoglobin.

7. Hemoglobin = 2a and 2b subunits
Labeling of a and b subunits:
3D Hemoglobin:

8. O2 Delivery by Hemoglobin
Your Hemoglobin delivers 45% more O2 when you exercise then at rest.
Hemoglobins ability to delivery 66% of it O2 to oxygen deprived cells makes it far superior to Myglobin for O2 delivery.

9. Grog the alien (often mistaken for old chemistry professors)
Grog the Alien hails from the Planet Weeiirddoo. Looking at Grogs Hb plot, would you think Weeiirddoo has more or less O2 than Earth?
Human Hb
Grog Hb
Grog the alien (often mistaken for old chemistry professors)
More O2 … as it requires more O2 to saturate his Hb

10. Hemoglobin: Conformation Changes with Binding of O2

11. Binding of O2 to the a-Subunit of Hemoglobin
Once O2 binds, Fe goes completely in plane with the heme.
In Deoxyhemoglobin, the iron atom is 0.4 angstroms out of plane
Fe movement pulls the bound His upward completely shifting an a-helix

12. 2,3-Bisphosphoglycerate Effect on Hemoglobin
The presence of BPG makes a difference of transferring 8% vs. 66% of the oxygen
Notice how this pocket is full of positively charged residues like Lys and His. This increases interaction with negatively charged BPG.
BPG binding pocket

13. BPG Binding Site in Embryos
Embryos, have different Hemoglobin that consists of 2a and 2g units. g is really similar to b, with the exception that at the BPG binding site His 143 is replaced w/Serines.
The result is BPG doesn’t bind as well and the T-state not as stable. This allows easy transfer of O2 from mothers hemoglobin to the babies hemoglobin.

14. Carbon Monoxide Poisoning
Due to similar shape, CO binds like O2 to Hemoglobin:
Why CO is deadly:
Binds Fe x stronger than O2 (out competes sites on Hb)
Forces Hb into R-state (BPG can’t bind and O2 bound can’t be released well)
The result of above is the suffocation of brain cells that leads to death
Roughly 2500 people die of CO each year: 2000 are suicides Treatment: expose to pure O2

15. His 146 and Asp 94 in Hemoglobin
At pH 7.2, about 33% of His 146 is protonated
At pH 7.4, about 20% of His 146 is protonated
Salt Bridges at Lower pH promote the T-state (which releases O2 to form deoxyhemoglobin)
Loss of Salt Bridges at Higher pH promote the R-state (which binds O2 to form oxyhemoglobin)

16. Transfer of CO2 from Tissue to Lungs

17. Dissection of Hemoglobin
What function do the outside polar Amino Acid residues on the a-helices perform?
What function do the salt bridges perform?
Stabilize the T-form of Hb
What functions do the lactic acid and CO2 perform?
Make hemoglobin water soluble (inside Red Blood Cells)
What functions do the Iron atoms perform?
What function does the His attached to the Iron atom perform?
Bind Oxygen and in doing so cause a small shift into the plane of the porphorin
The acid of lactic acid and Carbonic Acid (which is formed from CO2), protonate residues that form the salt bridges. The salt bridges push Hemoglobin toward a T-state which promotes the release of O2
Shifts an a-helix which converts the whole hemoglobin structure from a T-state to more of an R-state
What function does BPG perform?
Polar surface map of a-subunit in Hb
All red and blue are charged
Binds to a BPG pocket which stabilizes T-state, BPG pocket collaspes apon binding of 1st O2 to Hemoglobin

18. Sickle and Hammer in USSR symbol to indicate “worker party”
Sickle Cell Anemia
Normal
Red Blood
Cell
Sickle and Hammer in USSR symbol to indicate “worker party”
Sickle
Red Blood
Cell
“Sickeled Cells” can easily clog blood vessels
Sickeled Cell Anemia gene commonly found in Africa and India

19. Hemoglobin Sickle (HbS)
S fibers in Red Blood Cells
Under electron microscope

20. Sickled RBC Formed Upon Release of O2
O2 release
O2 binding
Normal looking RBC
R-state HbS
R-state HbS
Sickled RBC
A person suffering from sickle cell anemia will contain both “normal” looking red blood cells and sickled red blood cells. The “normal” looking cell will contain more R-state HbS than the sickled cells.

21. Sickle Cell and Malaria Density Maps

22. Woolly Mammoth’s Hemoglobin
Hemoglobin from a 43,000 yr old frozen baby woolly Mammoth extracted: Oldest know hemoglobin
Problem: Hemoglobin does not bind O2 from cold air. Since Woolly Mammoths lived in the arctic they had to adapt their hemoglobin to match the environment.

23. Wooly Mammoth’s Hemoglobin
Key Mutations (only on b chain, a chain contains no mutations): Thr12 -> Ala12 ; Ala86 -> Ser 86; Glu101 -> Gln 101
Glu 101 -> Gln 101 removes a salt bridge, reducing T-state Hb formation
Woolly Mammoth’s
Hb near the lungs
Normal Hb
Results of Mutation:
1. Woolly Hb binds O2 stronger near the lungs (this compensates for the cold).
2. Woolly Hb is more effected by changes in acidity (this compensates for the higher affinity of Hb for O2)
Woolly Mammoth’s
Hb near tissue