Define hemoglobin and state its function
- Hemoglobin: abundant, erythrocyte (red blood cell) protein
- Function: to pick up oxygen at lungs (high oxygen partial pressure, hence, concentration) and deliver it to tissues (lower partial pressure, hence, concentration)
Arteries for the most part carry _______ ______ to ______. What is the exception?
- oxygenated blood to tissues
- Exception: pulmonary artery (deoxygenated blood)
Veins for the most part carry _______ ______. What is the exception?
- deoxygenated blood
- Exception: pulmonary vein carries oxygenated blood
______ carries oxygen at lungs and delivers it to tissues, ______ is responsible for storage of oxygen
Why have we evolved an elaborate oxygen transport system?
Where is oxygen needed the most?
- Oxygen is insoluble, and needs transport protein
- Oxygen is most needed as the last electron acceptor in the electron transport chain
What is the three-dimensional shape of hemoglobin and myoglobin, if it has more than one form, specify.
How many subunits does hemoglobin have, is it monomeric or multimeric?
- 4, hemoglobin is a tetramer
- it is multimeric, 2 α-subunits and 2 β-subunits
In myoglobin, each subunit has _____ α-helices (lettered A-G). Each subunit binds _____ oxygen molecule(s). One hemoglobin molecule has _____ subunits. Thus, one hemoglobin molecule can bind ______ oxygen molecules
How many a-helices are present in myoglobin?
Each hemoglobin molecule (α1, α2, β1, β2) has a ______ heme group; _______ is monomeric. _______ groups are tightly bound (may or may not be covalently linked to protein)
- prosthetic heme group
- Prosthetic groups
What are the 5 functional groups in hemoglobin (memorize their positions and specific structures NOT hemoglobin's structure)
What are the red arrows pointing to? How many does Iron have? What is number 6 and number 5?
- Each red arrow points to a coordination center.
- Iron can have a total of 6
- #6 is the distal Histidine
- #5 is the proximal Histidine
In the absence of oxygen, what happens to iron? What happens upon oxygenation? What happens to the proximal histidine and the F-helix?
- The iron bulges out of the ring without oxygen and gets pulled back into the ring upon oxygenation, causing the dome-like shape to disappear/collapse
- The proximal histidine is pulled in as well
- Movement of F-helix (occurs in EACH subunit)
The nitrogen of the proximal histidine's imidazole makes a _______ linkage with the iron ion. Explain your answer
- Electronegativity Chart
- Fe = 1.83
- N = 3.04
- The difference is 1.21, hence a covalent bond
When does hemoglobin exhibit positive cooperativity? What is the defining characteristic, and what about it makes it positive cooperativity
- Hemoglobin exhibits positive cooperativity when binding to oxygen.
- Multimeric, ligand binding
- One subunit influences the other subunits to bind to ligand
Describe the hemoglobin and myoglobin dissociation curve
- Myglobin: hyperbolic curve, indicative of single subunit
- Hemoglobin: Sigmoidal, S-shaped curve, indicative of multiple subunits.
T-state (______) has a _____ binding state.
Inflection indicative of weak to strong transition is a representation of ___ (_______) to ____ (_______) transition
R state (_______) has a _____ binding state
- weak binding state
- T (deoxygenated) to R (oxygenated)
- strong binding state
________ releases most of its oxygen at pO2 levels at tissues BUT ______ does not
What causes low pH in the body?
Metabolically active cells releasing CO2
How is carbon dioxide generated in our bodies? Show the breakdown from molecule to molecule
Results from glucose catabolism (or oxidative breakdown in glycolysis)
Glucose →→→ Pyruvate → CO2 + Acetyl CoA
What is the major pathway through which carbon dioxide is removed from the body?
State the reaction
Major CO2 removal strategy: Enzyme catalyzing reaction generating Carbonic anhydrase
- H2O (l) + CO2 (g) ⇄ H2CO3(aq) ⇄ H+1 (aq) +HCO3-1 (aq)
What are two results of proton generation in the major CO2 removal pathway
- The protons lower our blood pH
- The protons bind to hemoglobin stabilizing the T state, which allows FASTER oxygen relase
What is the minor carbon dioxide removal strategy
It involves hemoglobin amino terminal carbamation
The T (_____, ________) state of hemoglobin is stabilized in two major ways (name them)
- Salt bridge formation
- BPG binding
Where do salt bridges form within a hemoglobin tetramer? How do they stabilize hemoglobin?
Salt bridges are formed between ends of a polypeptide chain AND between oppositely charged side chains
Helps hemoglobin molecule transition from a relaxed (R, oxygenated) to T (tense, deoxygenated) state
How many types of salt bridges can form in hemoglobin?
- 8 distinct salt bridges form
What is an allosteric effector?
Molecule that binds to a site on protein through non-covalent (hydrogen bonding, electrostatic/ionic) interactions
Which molecule does allosteric regulation of hemoglobin? What is the importance of this allosteric effector?
2,3-bisphosphoglycerate (BPG: a positive allosteric effector)
- 2,3-BPG stabilizes the T-state
How does 2,3 BPG bind? Give 4 characteristics
- Binds only to deoxyhemoglobin (2,3-BPG is big)
- Binds between the four subunits
- Binding mediated through favorable electrostatics, hydrogen bonding
- Upon oxygenation, 2,3-BPG is released/pushed out
What is the result of 2,3 BPG binding? What are the contributing residues?
The center of hemoglobin widens as a function of oxygen release stabilizing the T state (deoxygenated) and the R state (oxygenated) is effectively blocked
The contributing residues are Histidine and Lysine
How is 2,3-BPG produced?
- During glycolysis, 1,3-bisphosphoglycerate (a glycolytic intermediate) is produced
- Some 1,3-bisphosphoglycerate is diverted make an isomer known as 2,3-BPG
- It is isomerized via an enzyme knonw as 1,3-BPG mutase
How do 2,3-BPG concentration increase?
- 2,3-BPG is an isomer of the glycolytic intermediate 1,3-BPG which is produced during glucose breakdown
- 1,3-BPG increases during glycolysis hence, more can be isomerized to 2,3-BPG, which is an allosteric effector
This graph shows _______ curves of oxygen binding and release as a function of _____ _____.
What is the mMBPG and temperature at pH:
What activity would be associated with such levels?
- sigmoidal curves
- BPG concentration
- pH 7.20: 8 mM BPG and 40.0 °C (exercising making CO2)
- pH 7.40: 5 mM BPG and 37.4 °C (resting)
Explain protein allostery
When a small molecule makes its way into a groove or a pocket within a protein resulting in a positive or negative effect on protein function
Increase in BPG and temp. means _______ in pH
Decrease in BPG and temp. means _______ in pH
State 6 differences between the T state and R state
Two models work together to help describe hemoglobin's ability to bind oxygen. What are these models?
- When ONE out of FOUR subunits are bound with oxygen, hemoglobin follows a SEQUENTIAL model
- When THREE out of the FOUR subunits are bound with oxygen, hemoglobin follows a CONCERTED model
What do the subsequential and concerted model allow for?
What does this mean for the binding of each oxygen molecule?
Explain the oxygen binding affinity depicted at each arrow:
- A sequential model allows for the subsequent conformational change to each subunit in a multimeric protein
- In a concerted model, a substancial conformational change has ALREADY occurred to the protein (the change of the last molecule is minor)
- This means the binding of the FIRST THREE oxygen molecules to hemoglobin is sequential
- The binding of the LAST oxygen molecule is concerted