258
7 Respiratory Pigments
Glu [131, 132]. Xenopus haemoglobin contains
both ~FG1-Glu and ~F9-Ser and, despite previous findings to the contrary, still shows no Root
effect [26]. The haemoglobin from Lepidosiren
has the same amino acid composition and also
exhibits no Root effect. Thus, although it is possible to assign certain amino acids to key positions
in the haemoglobin molecule [131, 132], the functionally important properties of the haemoglobin
are nevertheless determined by the structure of
the whole molecule [21].
The goldfish possesses a single haemoglobin
showing a marked Root effect. The three principle haemoglobins of the carp are also Root-effect
haemoglobins. During rapid swimming, the
reduction in blood pH in the gill may be so great
that Root-effect haemoglobins can no longer be
saturated; in such cases, haemoglobins with and
without the Root effect are found together. The
best-known example of this is the rainbow trout
Salmo irideus (S. gairdneri): Hb IV (about 65 %
of the total) is a typical Root-effect haemoglobin,
but Hb I and II, in contrast, show practically no
Bohr effect; the properties of Hb III (about 3 %
of the total) are not known. In Hb I, all the polar
amino acids involved in the Bohr effect and in
binding organophosphates have been replaced by
neutral residues: ~EF6-Lys with leucine, ~F9-Ser
with alanine, ~FG1-Glu with asparagine, ~H21Arg with serine, and ~HC3-His with phenylalanine (Table 7.3). The sequence difference
between Hb IV and Hb I of the trout is no less
than 41 %, and between Hb IV and human Hb A
it only 50 %. This clearly illustrates the extreme
specialization of the Root-effect haemoglobins
[21, 131, 133]. Haemoglobin systems with mixed
properties, which provide adaptation to different
living conditions, are also found in adult amphibians. Whilst in the aquatic Surinam toad Pipa carvalhoi both principal haemoglobins show minor
Bohr effects ( with organophosphates, the related semi-aquatic
species Leptodactylus labrynthicus has the
"aquatic" Hb II and a further haemoglobin,
Hb III, which, like the haemoglobins of the terrestrial anurans, has a normal Bohr effect and a low
ATP sensitivity [107].
Carbon dioxide reduces the oxygen affinity
mainly by the protons it generates. A specific CO2
effect involves the formation of carbamino
groups with the terminal -NH2 of the a- and ~chains and a consequent reduction in O2 affinity.
Quantitatively, this effect plays only a minor role
compared with that of the H+ ions. Organophosphates compete with CO2 for the terminal-NH2 of
the ~-chains and thereby further reduce the CO2
effect. Blocking of the a-N-termini, as found in
the haemoglobins of many fish and amphibians
and of several mammals (Table 7.3), also prevents carbamino binding [183].
Certain amino acids of the ~-chains form positively charged fields after protonation and these
promote the T ~ R transition by mutual repulsion. In the mammals, this involves the Nterminus, ~NA2-His, ~EF6-Lys and ~H21-His.
Inorganic anions neutralize these charges and
thereby reduce the O2 affinity of most haemoglobins, although to a somewhat lesser degree than
do organophosphates. Because of its relatively
high concentration, the Cl- ion is of particular
physiological importance [24, 49]. The haemoglobin of the crocodile Caimand crocodylus is only
slightly sensitive to H+, CO2, Ct and organophosphates, and this clearly increases its diving capacity; conversely, HC03- has a reducing effect on O2
affinity, as in other crocodile species, and this
facilitates the efficient use of the oxygen bound to
the haemoglobin. The HC03- effect requires the
three amino acid substitutions ~NA1-Val to Ser,
~NA2-His to Pro and ~HC1-Lys to GIu; reduction
of the other heterotropic interactions comes from
the exchanges ~H21-His to Ala and aH14-Ser to
Ala [131, 183]. Urea, which in human Hb A increases the O2 affinity by dissociating the tetramers,
has no effect in several shark species, even at concentrations as high as 5.0 molll. The haemoglobins
of Squalus acanthias and Cephaloscyllium isabella
are, however, sensitive to urea: in contrast to various enzymes, the urea effect on haemoglobin is
not compensated by trimethylaminoxide [4].
Organophosphates like 2,3-bisphosphoglycerate (DPG) , inositol phosphates (IP6, IPs, etc.),
ATP or GTP induce salt bridges between the ~chains and thereby stabilize the Z state. In mammals DPG is bound to ~NA1-Val, ~NA2-His,
~EF6-Lys and ~H21-His, and in birds IP5 additionally binds to ~H13-Arg and ~H17-His. The teleosts have the amino acids ~NA2-Glul Asp and
~H21-Arg instead of the histidine found in the
equivalent positions in man; this represents a
steric adaptation to the binding of nucleoside triphosphates. ATP binds to five amino acids
(~2NA1-Val, ~lNA2-GIu, ~1H21-Arg, ~1+2EF6_
Lys); GTP additionally binds to ~lNA1-Val and
thereby has a twofold higher modulating effect
[183]. Curiously, the two haemoglobins of the rhinoceros Rhinoceros unicorn is contain not only the
DPG-binding sites typical for mammals, ~NA1Val, ~EF6-Lys and ~H21-His, but also in position
~NA2 either glutamic acid (Hb A) or aspartic acid
7 Respiratory Pigments
Glu [131, 132]. Xenopus haemoglobin contains
both ~FG1-Glu and ~F9-Ser and, despite previous findings to the contrary, still shows no Root
effect [26]. The haemoglobin from Lepidosiren
has the same amino acid composition and also
exhibits no Root effect. Thus, although it is possible to assign certain amino acids to key positions
in the haemoglobin molecule [131, 132], the functionally important properties of the haemoglobin
are nevertheless determined by the structure of
the whole molecule [21].
The goldfish possesses a single haemoglobin
showing a marked Root effect. The three principle haemoglobins of the carp are also Root-effect
haemoglobins. During rapid swimming, the
reduction in blood pH in the gill may be so great
that Root-effect haemoglobins can no longer be
saturated; in such cases, haemoglobins with and
without the Root effect are found together. The
best-known example of this is the rainbow trout
Salmo irideus (S. gairdneri): Hb IV (about 65 %
of the total) is a typical Root-effect haemoglobin,
but Hb I and II, in contrast, show practically no
Bohr effect; the properties of Hb III (about 3 %
of the total) are not known. In Hb I, all the polar
amino acids involved in the Bohr effect and in
binding organophosphates have been replaced by
neutral residues: ~EF6-Lys with leucine, ~F9-Ser
with alanine, ~FG1-Glu with asparagine, ~H21Arg with serine, and ~HC3-His with phenylalanine (Table 7.3). The sequence difference
between Hb IV and Hb I of the trout is no less
than 41 %, and between Hb IV and human Hb A
it only 50 %. This clearly illustrates the extreme
specialization of the Root-effect haemoglobins
[21, 131, 133]. Haemoglobin systems with mixed
properties, which provide adaptation to different
living conditions, are also found in adult amphibians. Whilst in the aquatic Surinam toad Pipa carvalhoi both principal haemoglobins show minor
Bohr effects ( with organophosphates, the related semi-aquatic
species Leptodactylus labrynthicus has the
"aquatic" Hb II and a further haemoglobin,
Hb III, which, like the haemoglobins of the terrestrial anurans, has a normal Bohr effect and a low
ATP sensitivity [107].
Carbon dioxide reduces the oxygen affinity
mainly by the protons it generates. A specific CO2
effect involves the formation of carbamino
groups with the terminal -NH2 of the a- and ~chains and a consequent reduction in O2 affinity.
Quantitatively, this effect plays only a minor role
compared with that of the H+ ions. Organophosphates compete with CO2 for the terminal-NH2 of
the ~-chains and thereby further reduce the CO2
effect. Blocking of the a-N-termini, as found in
the haemoglobins of many fish and amphibians
and of several mammals (Table 7.3), also prevents carbamino binding [183].
Certain amino acids of the ~-chains form positively charged fields after protonation and these
promote the T ~ R transition by mutual repulsion. In the mammals, this involves the Nterminus, ~NA2-His, ~EF6-Lys and ~H21-His.
Inorganic anions neutralize these charges and
thereby reduce the O2 affinity of most haemoglobins, although to a somewhat lesser degree than
do organophosphates. Because of its relatively
high concentration, the Cl- ion is of particular
physiological importance [24, 49]. The haemoglobin of the crocodile Caimand crocodylus is only
slightly sensitive to H+, CO2, Ct and organophosphates, and this clearly increases its diving capacity; conversely, HC03- has a reducing effect on O2
affinity, as in other crocodile species, and this
facilitates the efficient use of the oxygen bound to
the haemoglobin. The HC03- effect requires the
three amino acid substitutions ~NA1-Val to Ser,
~NA2-His to Pro and ~HC1-Lys to GIu; reduction
of the other heterotropic interactions comes from
the exchanges ~H21-His to Ala and aH14-Ser to
Ala [131, 183]. Urea, which in human Hb A increases the O2 affinity by dissociating the tetramers,
has no effect in several shark species, even at concentrations as high as 5.0 molll. The haemoglobins
of Squalus acanthias and Cephaloscyllium isabella
are, however, sensitive to urea: in contrast to various enzymes, the urea effect on haemoglobin is
not compensated by trimethylaminoxide [4].
Organophosphates like 2,3-bisphosphoglycerate (DPG) , inositol phosphates (IP6, IPs, etc.),
ATP or GTP induce salt bridges between the ~chains and thereby stabilize the Z state. In mammals DPG is bound to ~NA1-Val, ~NA2-His,
~EF6-Lys and ~H21-His, and in birds IP5 additionally binds to ~H13-Arg and ~H17-His. The teleosts have the amino acids ~NA2-Glul Asp and
~H21-Arg instead of the histidine found in the
equivalent positions in man; this represents a
steric adaptation to the binding of nucleoside triphosphates. ATP binds to five amino acids
(~2NA1-Val, ~lNA2-GIu, ~1H21-Arg, ~1+2EF6_
Lys); GTP additionally binds to ~lNA1-Val and
thereby has a twofold higher modulating effect
[183]. Curiously, the two haemoglobins of the rhinoceros Rhinoceros unicorn is contain not only the
DPG-binding sites typical for mammals, ~NA1Val, ~EF6-Lys and ~H21-His, but also in position
~NA2 either glutamic acid (Hb A) or aspartic acid