230
AUSTEN RIGGS
oxy and deoxy hemoglobin. TWO kinds of acid group on the protein are
functionally linked to oxygenation: one of these becomes more acidic
upon oxygenation (alkaline, normal Bohr effect, range p H 6.5-7.5),
while the other becomes less acidic (acid or “reverse” Bohr effect, range
pH 5.5-6.5). The alkaline Bohr effect is usually the only one which is
physiologically important. Although all the groups responsible for these socalled “Bohr” protons have not been unequivocally identified, the
acid Bohr effect may involve a carboxyl group and the alkaline Bohr
effect either imidazolc or the a-NH, group. The effective number of
protons released during oxygenation varies in different species; the direct
effect of CO, probably involves the formation of carbamino compounds :
-NH, + CO, + -NHCOOH. Recent evidence suggests that CO, is
bound primarily to the a-NH2 group of each chain in horse hemoglobin
(Kilmartin and Rossi-Bernardi, 1969) and that thc C terminal histidyl
residue of each , 8 chain is responsible for a major part of the alkaline
Bohr effect (Perutz et al., 1969); the a-NH, groups of the CY chains
are believed also to contribute to the Bohr effect.
The number of protons released during oxygenation can be estimated
from the dependence of the oxygen equilibrium on pH or directly by
differential titration. Wyman (1948, 1964) has shown that T [- ( 4 log
P 5 ” ) / A pH] equals the number of H+ released per oxygen bound provided that n is independent of p H (i.e., that the “y vs. log p’’ plots at
different values of p H are all parallel to one another). If, however, the
shape of the oxygen equilibrium curve changes with p H then the
median ligand activity (log p,,,) should be used rather than the ligand
activity associated with half-saturation (log P z 0 ) . The minimum change
in the pK of acid groups associated with oxygenation is given by the
expression ( Wyman, 1948) :
ApK = 2 10g[(l + r ) / ( l - r ) ]
Rossi-Fanelli and Antonini (1960) found that tuna hemoglobin has a
remarkably large Bohr effect but that the shape of the curve changed
with pH: the measured n drops from 3 to less than 1 when the pH drops
from 9 to 6. Indeed, at p H values lower than 6.5, complete saturation is
not achieved even at a partial pressure of one atmosphere of oxygen.
Brunori (1966) has studied the Bohr effect of this hemoglobin by the
differential titration method. He finds 0.86 H+ released pcr heme at pH
7.5 compared with 0.56 H+ for human hemoglobin. His results can be
fitted to a model for which he assumcs, as in human hemoglobin, only
two ligand-linked acid groups, one of which becomes more acidic
on oxygenation ( 4 p K E 2.2) and the othcr becomes lrss acidic ( A
pK
1.2). Only the first group is significant in the physiological range
AUSTEN RIGGS
oxy and deoxy hemoglobin. TWO kinds of acid group on the protein are
functionally linked to oxygenation: one of these becomes more acidic
upon oxygenation (alkaline, normal Bohr effect, range p H 6.5-7.5),
while the other becomes less acidic (acid or “reverse” Bohr effect, range
pH 5.5-6.5). The alkaline Bohr effect is usually the only one which is
physiologically important. Although all the groups responsible for these socalled “Bohr” protons have not been unequivocally identified, the
acid Bohr effect may involve a carboxyl group and the alkaline Bohr
effect either imidazolc or the a-NH, group. The effective number of
protons released during oxygenation varies in different species; the direct
effect of CO, probably involves the formation of carbamino compounds :
-NH, + CO, + -NHCOOH. Recent evidence suggests that CO, is
bound primarily to the a-NH2 group of each chain in horse hemoglobin
(Kilmartin and Rossi-Bernardi, 1969) and that thc C terminal histidyl
residue of each , 8 chain is responsible for a major part of the alkaline
Bohr effect (Perutz et al., 1969); the a-NH, groups of the CY chains
are believed also to contribute to the Bohr effect.
The number of protons released during oxygenation can be estimated
from the dependence of the oxygen equilibrium on pH or directly by
differential titration. Wyman (1948, 1964) has shown that T [- ( 4 log
P 5 ” ) / A pH] equals the number of H+ released per oxygen bound provided that n is independent of p H (i.e., that the “y vs. log p’’ plots at
different values of p H are all parallel to one another). If, however, the
shape of the oxygen equilibrium curve changes with p H then the
median ligand activity (log p,,,) should be used rather than the ligand
activity associated with half-saturation (log P z 0 ) . The minimum change
in the pK of acid groups associated with oxygenation is given by the
expression ( Wyman, 1948) :
ApK = 2 10g[(l + r ) / ( l - r ) ]
Rossi-Fanelli and Antonini (1960) found that tuna hemoglobin has a
remarkably large Bohr effect but that the shape of the curve changed
with pH: the measured n drops from 3 to less than 1 when the pH drops
from 9 to 6. Indeed, at p H values lower than 6.5, complete saturation is
not achieved even at a partial pressure of one atmosphere of oxygen.
Brunori (1966) has studied the Bohr effect of this hemoglobin by the
differential titration method. He finds 0.86 H+ released pcr heme at pH
7.5 compared with 0.56 H+ for human hemoglobin. His results can be
fitted to a model for which he assumcs, as in human hemoglobin, only
two ligand-linked acid groups, one of which becomes more acidic
on oxygenation ( 4 p K E 2.2) and the othcr becomes lrss acidic ( A
pK
1.2). Only the first group is significant in the physiological range