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

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.

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.

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.

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

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.

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

Hemoglobin: Conformation Changes with Binding of O2

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

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

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.

Carbon Monoxide Poisoning Due to similar shape, CO binds like O2 to Hemoglobin: Why CO is deadly: Binds Fe2+ 1000x 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

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)

Transfer of CO2 from Tissue to Lungs

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

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

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

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.

Sickle Cell and Malaria Density Maps

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.

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