Biology Reference
In-Depth Information
effect, however, because of the inadequacy of available experimental
techniques, and thus concluded incorrectly that O
2
did not affect CO
2
binding. Again, remember these experiments were carried out more than
100 years ago.
Bohr's steadfast belief in the experimental approach probably prevented
him from seeing what now seems an obvious inference; namely, that
the reciprocal effects of the binding of O
2
and CO
2
by blood must
exist. Clearly, if CO
2
affects the binding of O
2
, then O
2
must affect the
binding of CO
2
. It appears Bohr trusted his data too much, while
H ¨ fner trusted his theoretical model too much. Evidently, both
experiments and theory are required, and neither should be favored to
the exclusion of the other.
C. The Hill Equation
The apparent inconsistency between H ¨ fner's theory of noncooperative
binding, Eqs. (7-6) and (7-7), and Bohr's experimentally observed
cooperative O
2
binding data was difficult to resolve. Both seemed to be
correct. Then in 1910, A. V. Hill (1886-1977; Nobel laureate, 1929)
developed a conceptual model that appeared to reconcile experiment
and theory. Hill realized H ¨ fner's theory would be correct if hemoglobin
contained only a single O
2
binding site, but the sigmoid-shaped O
2
binding data of Bohr contradicted this theory, and, therefore,
hemoglobin must have more than one binding site. Hill, however, did
not know how many binding sites there were—his research predated the
concept of unique multi-subunit high-molecular weight protein
molecules by two decades. At this time, proteins were thought to be
heterogeneous aggregates. Hill postulated that aggregates with
n monomers would bind n molecules of O
2
according to a reaction like:
Hb
n
þ
nO
2
$
Hb
n
ð
O
2
Þ
n
;
(7-8)
where Hb
n
is used to denote the assumption made by Hill that
hemoglobin is constructed of n monomers. Hill did not know what the
actual value of n was and wanted to determine the value experimentally.
Hill's scheme assumes that n molecules of O
2
are bound simultaneously
in a single step. This is equivalent to assuming no intermediate states
ever exist where the number of O
2
molecules bound is greater than
zero and less than n. Given this assumption and the assumption that
only one aggregate (i.e., a single value of n) exists, Hill formulated an
equation for the fractional saturation as:
n
k
½
O
2
Y
¼
:
(7-9)
n
1
þ
k
½
O
2
This Hill equation is still in use today. The n is the Hill coefficient that is
sometimes used as a measure of cooperativity. For n
1, Eqs. (7-8)
and (7-9) predict a sigmoid-shaped binding curve like the H curve in
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