8 - Haemoglobin and Cooperativity Flashcards
What is the structure of haemoglobin?
Four helical subunits - two alpha and two beta. Each containing a haem group.
What is the structure of myoglobin?
A single subunit with a haem group, comparative to one of the haemoglobin subunits.
What is AHSP?
Alpha-haemoglobin Stabilising Protein - a chaperone which stabilises the alpha haemoglobin subunits prior to formation into the haemoglobin complex.
What is the structure of the haem group?
It is a prophyrin ring, comprising four pyrrole rings whose endocyclic nitrogens coordinate the iron atom.
What state is the haem iron in when not bound to oxygen?
Ferrous, +2
How many coordination bonds can the anoxygenic ferrous iron form?
Six.
How many of the coordinate bonds of the ferrous iron are fulfilled by the porphyrin ring?
Four
What part of the subunit binds to the fifth coordination site of the ferrous iron, and what effect does this have?
The proximal histidine on the F-helix, whose coordination pulls the iron downwards, puckering the ring.
How does oxygen reach its binding site?
The haem group is deeply buried, so the O2 only reaches it due to local breathing of the protein.
What occurs when the oxygen binds to the ferrous iron?
It pulls the iron back into plane with the porphyrin ring, undoing the puckering. It also partially donates an electron producing ferric (+3) iron and becoming a superoxide radical.
What binding event stabilises the superoxide radical?
Hydrogen bonding of the second oxygen atom with the distal histidine on the E-helix.
What are the forms of the oxygen affinity curves for myoglobin and haemoglobin?
Myoglobin - rectangular hyperbola
Haemoglobin - sigmoidal
What is the percentage change in oxygen affinity for myoglobin and haemoglobin between oxygen partial pressures at the tissues and lungs?
7% for myoglobin
66% for haemoglobin
What would the oxygen affinity curve look like for haemoglobin if it did not show cooperative binding?
Much higher Km than Hb, to the point where it does not plateau until well after the maximum physiclogical concentration. It has a 38% change in affinity over the difference between lung and tissue O2 pp.
What is the mechanism for haemoglobin cooperativity?
When oxygen binds to a subunit it causes a conformational change which effects a 15° rotation relative to the other subunits. It is this that increases the oxygen affinity of the adjacent chains.
What are the two models of three models of cooperativity?
Concerted
Sequential
Mnemonic
What is the concerted model of cooperativity in the context of haemoglobin?
The concerted model postulates that the haemoglobin subunits can either all be tense or all relaxed, and that oxygen binding to more subunits shifts the equilibrium towards the relaxed state.
What is the sequential model of cooperativity in the context of haemoglobin?
The sequential model assumes that the subunits can exist in different states within the same Hb protein, with oxygen binding to one of them converting it to the relaxed state which affects the likelihood of the adjacent subunits relaxing.
What is used to measure the cooperativity of the binding of a protein?
Its Hill Coefficient, derived from a Hill plot.
What coefficients are Hill plots derived from?
Hill equation requires logarithmic transformation of an equation involving number of binding sites, ligand concentration and Kd.
How is the Hill coefficient derived from a Hill plot?
It is the gradient of the line.
What does a Hill coefficient of 1 entail?
Lack of cooperativity.
What do high and low Hill coefficients describe?
> 1 = positive cooperativity
What are the Hill coefficients of myoglobin and haemoglobin?
Haemoglobin = 2.8
Myoglobin = 1
