Haemoglobin and Allostery Flashcards
haem
heam (heme) is a cofactor consisting of an Fe2+ (ferrous iron) ion contained in the centre of a large heterocyclic organic ring called a porphyrin, which may or may not be covalently attatched to protein backbone
how does haem spatially bind oxygen in a globin?
allostery
proteins that have multiple binding sites (either the same or different), in which the binding of the ligand at the first site (allosteric site) causes a conformational change that alters the affinity of binding at the second site.
positive or negative cooperativity, depending on whether the allosteric interaction promotes or inhibits binding at the second site
contrast the shape of the graph of θ vs. ligand concentration in cooperative binding and standard allostery
Cooperative binding leads to a sigmoidal binding curve rather than a hyperbolic curve found in standard allostery
describe the shape of myoglobin in a graph of θ vs. ligand concentration
• Myoglobin consists of a single globin chain, and therefore has a typical hyperbolic binding curve.
At partial pressures of oxygen typically found in both the lungs and tissues, it remains highly saturated with oxygen making it an effective oxygen storage molecule, particularly in muscle. If the oxygen level falls below the normal levels required by the tissues, then myoglobin begins releasing oxygen to supply the cell with its needs
describe the two states of any given subunit in haemoglobin
The subunits of Hb exist in 2 states;
- A “tense” or “T” state, to which it is harder for O2 to bind.
- A “relaxed” or “R” state, which facilitates O2 binding.
Binding of the first molecule of O2 to deoxygenated Hb causes conformational changes that make binding of O2 to the other subunits easier.
(shown: Changes in conformation near heme on O2 binding to deoxyhemoglobin. (Derived from PDB ID 1HGA and 1BBB) The shift in the position of helix F when heme binds O2 is thought to be one of the adjustments that triggers the T → R transition.)
describe the models of cooperative binding in haemoglobin
Two general models for the interconversion of inactive and active forms of a protein during cooperative ligand binding. Although the models may be applied to any protein—including any enzyme (Chapter 6)—that exhibits cooperative binding, we show here four subunits because the model was originally proposed for hemoglobin. (a) In the concerted, or all-or-none, model (MWC model), all subunits are postulated to be in the same conformation, either all (low affinity or inactive) or all (high affinity or active). Depending on the equilibrium, K1, between and forms, the binding of one or more ligand molecules (L) will pull the equilibrium toward the form. Subunits with bound L are shaded. (b) In the sequential model, each individual subunit can be in either the or form. A very large number of conformations is thus possible.
behaviour of haemoglobin is consistant with both models, exact mechanism still under study
sigmoidal curve
A sigmoidal (cooperative) binding curve can be viewed as a hybrid curve reflecting a transition from a low-affinity to a high-affinity state. Because of its cooperative binding, as manifested by a sigmoid binding curve, hemoglobin is more sensitive to the small differences in O2 concentration between the tissues and the lungs, allowing it to bind oxygen in the lungs (where pO2 is high) and release it in the tissues (where pO2 is low).
describe the alloseric transistion in haemoglobin
Considerable energy is required to bind the first molecule of O2 to a subunit in the T-state, which requires the breaking of ionic interactions between the subunits to open up the site. Once this occurs, and O2 remains bound, the T-state cannot reform.
• The conformational change primarily involves rotations between the subunits opening up the other heme binding sites, allowing further molecules of O2 to bind (concerted or “all of nothing” model). The full conformational changes converting the T-state to the R-state are not complete until all the heme sites are occupied, and hemoglobin is fully oxygenated (sequential model).
What role could the indicated amino acid side chains play in O2 binding in haem?
His E7 hydrogen bonds with O2, stabilising the structure.
Val E11 is involved in regulating ligand recombination
DeoxyHb
haemoglobin in the T-state, with the haem in the Fe2+ state.
OxyHb
haemoglobin in the R-state, with all 4 sites occupied by molecular oxygen bound to Fe2+ (ferrious iron).
which state of haemoglobin is more stable in the presence of O2?
the R-state, the Fe2+-O2 bond is more exergonic (in a lower energy state) in the R-state than in the T-state.
R-state has a higher affinity for O2.
In the absence of O2, T-state is more stable; when O2 binds, R-state is more stable, so hemoglobin undergoes a conformational change to the R-state.
The structural change involves readjustment of interactions between subunits
which state of haemoglobin is more stable in the absence of O2?
the T-state, the Fe2+-O2 bond is more exergonic (in a lower energy state) in the R-state than in the T-state.
R state has a higher affinity for O2.
In the absence of O2, T-state is more stable; when O2 binds, R-state is more stable, so hemoglobin undergoes a conformational change to the R-state.
The structural change involves readjustment of interactions between subunits
describe the Bohr effect on haemoglobin
O2 binding to haemoglobin is dependent on pH and CO2
Lower pH and/or increased CO2 causes O2 to bind with less afinity
In tissues, where pH is relatively low and the CO2 concentration is relatively high (due to production by biochemical reactions in the tissues), haemoglobin “drops” O2, which is released to the tissues. Haemoglobin binds some excess CO2 and becomes protonated (binds H+)
When the blood cell reaches the lungs, the pH rises and some of the CO2 is expelled, lowering the concentration and allowing haemoglobin to bind O2 again.
BPG
D-2,3-bisphosphoglycerate is formed in red blood cells from a glycolytic intermediate, binds between subunits of haemoglobin making conversion from the T-state to the R-state more difficult, lowering Hb’s affinity for O2.
This is required to relase suitable amounts of O2 near tissues
D-2,3-bisphosphoglycerate
D-2,3-bisphosphoglycerate (BPG) is formed in red blood cells from a glycolytic intermediate, binds between subunits of haemoglobin making conversion from the T-state to the R-state more difficult, lowering Hb’s affinity for O2.
This is required to relase suitable amounts of O2 near tissues
what modification to the gene product causes sickle cell anemia?
Caused by a single point mutation on the β chain of hemoglobin from Glu6 to Val6.
The HbS (sickle variant) Val6 fits perfectly into a hydrophobic pocket of another Hb in the deoxygenated form (this pocket does not exist in oxyHb because of its R-state structure) forming strands, deforming the red blood cell.
describe the shape of haemoglobin in a graph of θ vs. ligand concentration
• Hemoglobin exhibits positive cooperativity, and the graph is sigmoidal.
At the lungs Hb is saturated with oxygen (it “picks up” oxygen) but at the levels normally found in tissues, it releases a significant amount (the lower the tissues’ levels, the more it releases) thus making it a good oxygen transport molecule.
Bohr effect
The Bohr effect is a physiological phenomenon first described in 1904 by the Danish physiologist Christian Bohr, stating that hemoglobin’s oxygen binding affinity is inversely related both to acidity and to the concentration of carbon dioxide. Id est, an increase in blood CO2 concentration which leads to a decrease in blood pH will result in hemoglobin proteins releasing their load of oxygen. Conversely, a decrease in carbon dioxide provokes an increase in pH, which results in hemoglobin picking up more oxygen. Since carbon dioxide reacts with water to form carbonic acid, an increase in CO2 results in a decrease in blood pH.
