Biology

Couple Case Study Questions (Sickle Cell Anemia&Crocodile Hemoglobin)

Allosteric Interactions in Crocodile Hemoglobin
Focus concept
The effect of allosteric modulators on oxygen affinity for crocodile hemoglobin is unique when
compared with other species.
Prerequisite
Hemoglobin structure and function concepts.
Background
While most human beings are able to hold their breath for only a minute or two, other species are able
to stay under water for much longer periods of time. In this case study we will examine the physiological
adaptations that allow some organisms to deliver oxygen to tissues while submerged under water.
Deep sea-diving mammals, such as whales and seals, are able to stay under water for long periods of
time. These mammals are able to stay submerged because their muscles contain many-fold higher
concentrations of myoglobin (Mb) than humans.
Crocodiles are also able to stay submerged under water for periods of time exceeding one hour. This
adaptation allows the crocodile to kill small mammals by drowning them. However, the crocodile doesn’t
have large amounts of myoglobin in its muscle as the deep sea-diving mammals do, so their physiological
adaptation must be different. In 1995, Nagai and colleagues described in the British journal Nature a
possible mechanism that allowed the crocodile hemoglobin to deliver a large fraction of bound oxygen to
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the tissues. They suggested that bicarbonate, HCO , binds to hemoglobin to promote the dissociation of –
oxygen in a manner similar to 2,3-bisphosphoglycerate (BPG) in humans.
This case is important because information gathered from experiments like those described here will
allow scientists to design effective blood replacements.
Questions
1. In humans, oxygen is effectively delivered to the tissues because of the presence of several allosteric
modulators. Name three of these modulators and explain how their presence allows oxygen to be
delivered to the tissues.
2. Explain why having higher concentrations of Mb would allow whales and seals to stay submerged
under water for a long period of time.
3. Let us consider the hypothesis that bicarbonate serves as an allosteric modulator of hemoglobin
b 3
inding in crocodiles. What is the source of HCO in the crocodile tissues? –
CASE 9 C Allosteric Interactions in Crocodile Hemoglobin
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4. Draw oxygen-binding curves for crocodile hemoglobin in the presence and absence of bicarbonate.
Wh 50 ich conditions give rise to a greater p value for crocodile hemoglobin? What does this tell you
about the oxygen binding affinity for hemoglobin under those conditions?
5. Komiyama et al. investigated the bicarbonate binding site on the crocodile hemoglobin by
constructing human-crocodile chimeric hemoglobins in which amino acids in the human hemoglobin
were replaced with amino acids found in the crocodile hemoglobin at the same location. (The
investigators wanted to see if they could make a synthetic human hemoglobin that resembled the
crocodile hemoglobin in terms of its ability to bind bicarbonate anions.) They found the bicarbonate
b 1 2 inding site to be located at the ” $ -subunit interface, where the two subunits slide with respect to
one another during R W T transitions. Based on their results the authors modeled a stereochemically
plausible binding site that included the phenolate anion of Tyr 41$, the ,-amino group of Lys 38$,
and the phenolate anion of Tyr 42″.
What kinds of interactions do you think the aforementioned amino acid side chains will have with the
bicarbonate anion? (It might be helpful to draw the Lewis electron dot structure of bicarbonate).
6. In order to create an engineered human hemoglobin molecule that had the same bicarbonate binding
properties as crocodile hemoglobin, twelve amino acid residues had to be changed. Not all of these
residues directly interact with bicarbonate–perhaps only three of them do, as described in question 5.
What might be the role of the other nine amino acid residues?
7. Other animals have similarly adapted to using small molecules as allosteric effectors to encourage
hemoglob 3
in to release its oxygen. Whereas humans use 2,3-BPG and crocodiles use HCO , birds use –
myo 5 5 -inositol pentaphosphate (IP ) and fish use ATP and GTP. The structures of ATP and IP are
shown in Figure 9.1. What structural characteristics do all of these molecules have in common and
how would they bind to hemoglobin?
Reference
Komiyama, N. H., Miyazaki, G., Tame, J., and Nagai, K. (1995) Nature 373, pp. 244-246.
CASE 9 C Allosteric Interactions in Crocodile Hemoglobin
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Figure 9.1: Allosteric effectors of hemoglobin in various species.

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