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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)
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Arteries for the most part carry _______ ______ to ______. What is the exception?
- oxygenated blood to tissues
- Exception: pulmonary artery (deoxygenated blood)
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Veins for the most part carry _______ ______. What is the exception?
- deoxygenated blood
- Exception: pulmonary vein carries oxygenated blood
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______ carries oxygen at lungs and delivers it to tissues, ______ is responsible for storage of oxygen
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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
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What is the three-dimensional shape of hemoglobin and myoglobin, if it has more than one form, specify.
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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
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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
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How many a-helices are present in myoglobin?
8
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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
- myoglobin
- Prosthetic groups
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What are the 5 functional groups in hemoglobin (memorize their positions and specific structures NOT hemoglobin's structure)
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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
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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)
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The nitrogen of the proximal histidine's imidazole makes a _______ linkage with the iron ion. Explain your answer
covalent
- Electronegativity Chart
- Fe = 1.83
- N = 3.04
- The difference is 1.21, hence a covalent bond
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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
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Describe the hemoglobin and myoglobin dissociation curve
- Myglobin: hyperbolic curve, indicative of single subunit
- Hemoglobin: Sigmoidal, S-shaped curve, indicative of multiple subunits.
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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
- deoxygenated
- weak binding state
- T (deoxygenated) to R (oxygenated)
- oxygenated
- strong binding state
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________ releases most of its oxygen at pO2 levels at tissues BUT ______ does not
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What causes low pH in the body?
Metabolically active cells releasing CO2
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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
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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
- Reaction:
- H2O (l) + CO2 (g) ⇄ H2CO3(aq) ⇄ H+1 (aq) +HCO3-1 (aq)
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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
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What is the minor carbon dioxide removal strategy
It involves hemoglobin amino terminal carbamation
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The T (_____, ________) state of hemoglobin is stabilized in two major ways (name them)
tense, deoxygenated
- Salt bridge formation
- BPG binding
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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
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How many types of salt bridges can form in hemoglobin?
- 8 distinct salt bridges form
 
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What is an allosteric effector?
Molecule that binds to a site on protein through non-covalent (hydrogen bonding, electrostatic/ionic) interactions
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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
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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
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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
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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
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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
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This graph shows _______ curves of oxygen binding and release as a function of _____ _____.
What is the mMBPG and temperature at pH:
7.2
7.4
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)
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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
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Increase in BPG and temp. means _______ in pH
Decrease in BPG and temp. means _______ in pH
- decrease
- increase
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State 6 differences between the T state and R state
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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
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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)
- Meaning:
- This means the binding of the FIRST THREE oxygen molecules to hemoglobin is sequential
- The binding of the LAST oxygen molecule is concerted
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