lecture 8 - allostery & cooperativity in haemoglobin Flashcards
What is a conformational change of a protein?
A change in the shape of secondary elements of a protein
What is the shape of the haem unit in deoxyhaemoglobin?
Dished
What is the shape of haem in oxyhemoglobin?
Flattened
How does O2 cause a conformational change in Haem binding to Hb?
O2 flattens the Haem, pulling Fe2+ into plane, and pulls His F8 of helix F towards the binding site.
How does proximity of haem and Helix F affect Hb O2 binding?
Anything that keeps helix F away from the haem unit weeks oxygen binding
What helices are involved in alternative side chain packing between the T and R states in Hb?
the F and C helices of adjacent subunits. (e.g. alpha1 and Beta2)
What is one of the key differences in bonding/side chain packing in the F and C helices of adjacent subunits between the oxygenated (R) and deoxygenated (T) states?
Tyrosine residue on C helix moves from being H-bonded to aspartic acid on F helix to being unbonded
What is the unabbreviated name for BPG?
2,3-biphosphoglycerate
Where does BPG originate, and what is its fucntion?
A molecule made during metabolism that stabilises the T-state of HB by binding to an allosteric site
What is the charge on BPG?
5 negative charges
What is the charge on the allosteric site of Hb that BPG binds to, and why?
Positive, due to BPG’s 5 negative charges, allowing for electrostatic interaction/binding
What is the physiological function of BPG?
Made during respiration, and binds to deoxy-Hb in T state, stabilising it and reducing oxygen affinity so O2 can be delivered to peripheral tissues.
What are the 3 key allosteric inhibitors that influence Hb binding and cooperativity?
BPG, CO2, H+
What is the effect of allosteric inhibitors on Hb?
Stabilise the T state, reducing oxygen affinity
In the absence of inhibitors, what state is Hb predominantly in?
The R-state, because the inhibitors stabilise the T-state
What are the 2 ways that CO2 reduced O2 affinity of Hb?
Directly, and via lowering the pH of blood
What is the Bohr effect?
Decrease in the affinity of Hb to oxygen due to increased CO2 levels and low pH of the blood.
Where does CO2 bind to to stabilise the deoxy-Hb conformation in the T state?
Extreme N-terminal amino groups
Where does H+ bind to to stabilise the deoxy-Hb conformation in the T state?
H+ protonates certain amino side chains
What is the reaction for the reversible binding of CO2 and H+ to Hb?
HbO2 + H+ + CO2 ⇌ O2 + Hb-CO2-H+ (CO2 and H+ bound to Hb)
What binding reaction involving CO2 and H+ occurs in actively metabolising tissue, such as muscle?
Oxygen unloading - HbO2 + H+ + CO2 ⇌ O2 + Hb-CO2-H+
What binding reaction involving CO2 and H+ occurs in the alveoli of the lungs?
Oxygen loading - O2 + Hb-CO2-H+ ⇌ HbO2 + H+ + CO2
What happens to BPG concentration at high altitude?
BPG production increases
What is the function of increased BPG concentration at high altitude?
Reduces Hb’s affinity to O2, allowing more oxygen to be delivered to tissues - a rightward shift of the binding curve
Does foetal haemoglobin have a higher or lower affinity for oxygen?
Higher
How does foetal haemoglobin differ to adult haemoglobin?
They have different isoforms/subunits with higher O2 affinity
What are the alternative subunits that foetal haemoglobin has?
ε (epsilon), γ (gamma), ζ (zeta)
What is the fucntion of the alternative subunits found in foetal Hb?
Increased affinity to O2 allows the foetus to capture O2 in the placenta
What is the relative sensitivity of foetal Hb to BPG?
Low sensitivity to BPG, so binds O2 tightly
What chain replaces the Beta chain of adult Hb in foetal Hb?
Gamma chain
What is the oxidation state of Fe in deoxy-Hb?
Fe2+
What is the oxidation state of Fe in oxy-Hb?
Fe2+
What is the oxidation state of Fe in methaemoglobin?
Fe3+
What happens to binding in methaemoglobin?
The oxidation of Fe2+ to Fe3+ shifts one subunit to the R-state conformation without O2 binding (as O2 cannot bid to Fe3+). Cooperativity means that the other subunits of the tetramer are shifted to the R state, so cannot release O2 to the tissues as they should
What enzyme is used to converted methaemoglobin back to haemoglobin?
cytochrome b5 reductase
How does cytochrome b5 reducatase regenerate methaemoglobin to haemoglobin?
Reduces the Fe3+ back to Fe2+, using electron transfer via the electron carrier NADH
What is the symbol for Boston haemoglobin?
HbM
What is the oxidation state of Fe in Boston Hb (HbM)?
Fe3+
How does HbM/Boston Hb function?
All subunits of HbM remains in the T state due to the Fe3+ Haem units (methaemoglobin), and oxygen cannot bind
What is the symbol for sickle cell haemoglobin?
HbS
What is the affect of sickle cell haemoglobin on red blood cells?
Red blood cells have an abnormal shape and get stuck in blood capillaries
What type of mutation causes sickle cell haemoglobin?
Gain of function mutation
What occurs in a HbS mutation?
Protein mutation enables an abnormal hydrophobic interaction between Hb molecules, particularly in their deoxy- form, causing polymerisation of HB into chains that distort the red blood cells
What part of haemoglobin does BPG bind to?
A positively charged pocket in the middle/core
