magna produces a complex spectrum of subunits
under both reducing and non-reducing conditions; these subunits apparently arise partially by
reciprocal conversion. The structure of these haemoglobins is thus more complicated than was originally assumed [130]. Crustacean haemoglobins
show only weak cooperativity (e.g. for Artemia
n= 1.6-1.9) and never a strong Bohr effect. Haemoglobin synthesis in Daphnia is stimulated by
reductions on pOz, and in Artemia salina by an
increase in the salt concentration of the environment. The haemoglobin concentration in the haemolymph of both species shows corresponding
wide fluctuation, reaching 15 gil in Daphnia and
up to 20 gil in Artemia salina. Curiously, the concentration in the Australian species Parartemia
zietziana is always very low (0.1 gil) and is not
influenced by the pOz [85].
7.2.4 Haemoglobins of the Insects
Intracellular haemoglobins are found in only a
few insect species. Larvae of the fly Gasterophilus
intestinalis, which live in the horse stomach, initially have haemoglobin in various body cells, but
in later developmental stages haemoglobin occurs
only in tracheal cells. The Gasterophilus haemoglobin is a non-cooperative homodimer of
34 kDa, and belongs to the few haemoglobins
which have a lower affinity for CO than for Oz.
Tracheal cell haemoglobin is also found in the
back-swimmers (Notonectidae) of the genera
Anisops and Buenoa. The haemoglobin of Anisops assimilis is a monomer of 16 kDa in the oxygenated state, and aggregates on deoxygenation.
The Hill plot is triphasic with n = 1.1, 5.2 and
2.6. The extremely high cooperativity in physiological conditions is explained entirely by the Ozdependent dissociation. In this case, the haemoglobin assists in the production of an Oz gas-bubble
for regulating buoyancy. Buenoa haemoglobin
also tends to aggregate. Intracellular haemoglobins have also been reported in the accessory
glands of the male genital tract of the water bug
Macrocorixa geoff roy and in the eggs of the louse
Pediculus humanus and the blood-sucking bug
Rhodnius prolixus [128, 174].
Extracellular haemoglobins are restricted to
the larvae of the non-biting midges (Chironomidae). These inhabit the mud regions of stagnant
water and appear red due to the haemoglobin dissolved in the haemolymph. As in the case of
Daphnia and Artemia, the haemoglobin concentration is higher in animals from low-Oz biotopes.
7.2.4 Haemoglobins of the Insects
273
The concentration in most species is highest during the fourth larval stage, and shows a marked
seasonal rhythm with a maximum in summer. In
extreme cases, haemoglobins make up more than
90 % of the haemolymph protein [104]. Contrary
to the rule that extracellular haemoglobins have
high molecular weights, the Chironomus haemoglobins are monomers or dimers, and seldom
tetramers (c. strenzkei) , of 16-kDa subunits.
Dimeric forms predominate in most species. All
the chironomid species possess 6-12 different
haemoglobins encoded by different genes. Twelve
haemoglobins have been described in the most
investigated species C. thummi thummi: these are
monomers CIT-I, -lA, -III, -IlIA, and -IV;
homodimers CTT-Il~, -VI, -VIlA, -VIlB, -VIII
and -IX; and CIT-X exists as monomers and
dimers in equilibrium [128].
The amino acid sequences are known for all
12 haemoglobins of C. thummi thummi [81, 108].
The chains vary in length between 136 and 151
amino acids due to extensions at both ends or to
deletions. Pairwise sequence comparisons give a
consistent 50 % agreement; 21 positions are
invariant if CIT-IlIA is not considered, but only
12 are conserved when this haemoglobin is
included in the analysis. Thus, all these haemoglobins apparently arose 200-400 million years
ago from a common monomeric ancestor. The
three-dimensional structure has been examined
only for CIT-III (Fig. 7.4) but, including all Hbridges, haem contacts etc., the analysis is at least
as thorough as for sperm whale myoglobin.
Dimer formation involves the interaction of G 19His (Lys in CTT-Il~) and G7-Glu. The monomeric forms contain G19-His but lack G7-Glu
[128]. The haem has the usual proximal contacts,
including F8-His, but the distal haem contacts are
unusual. With the exception of CIT-I, position
Ell is always occupied by isoleucine; this interacts with the haem iron and forces the E7-His out
of the haem pocket. CTT-I has valine at position
Ell and the Fe contact here is possibly via E7His. E7 in CIT-IlIA is occupied by glutamine.
The haemoglobins CIT-III and CTT-IV include
isomers in which the haem is rotated 180 0 , as has
already been described for vertebrate myoglobins
and the haemoglobin of the annelid Glycera
[31, 128].
It has been proposed that the site of haemoglobin biosynthesis in the chironomids lies not only
in the fat bodies but also in the oenocytes; however, tracer experiments have now shown that
haemoglobins are synthesized in subepithelial fat
bodies and temporarily stored in the oenocytes.
under both reducing and non-reducing conditions; these subunits apparently arise partially by
reciprocal conversion. The structure of these haemoglobins is thus more complicated than was originally assumed [130]. Crustacean haemoglobins
show only weak cooperativity (e.g. for Artemia
n= 1.6-1.9) and never a strong Bohr effect. Haemoglobin synthesis in Daphnia is stimulated by
reductions on pOz, and in Artemia salina by an
increase in the salt concentration of the environment. The haemoglobin concentration in the haemolymph of both species shows corresponding
wide fluctuation, reaching 15 gil in Daphnia and
up to 20 gil in Artemia salina. Curiously, the concentration in the Australian species Parartemia
zietziana is always very low (0.1 gil) and is not
influenced by the pOz [85].
7.2.4 Haemoglobins of the Insects
Intracellular haemoglobins are found in only a
few insect species. Larvae of the fly Gasterophilus
intestinalis, which live in the horse stomach, initially have haemoglobin in various body cells, but
in later developmental stages haemoglobin occurs
only in tracheal cells. The Gasterophilus haemoglobin is a non-cooperative homodimer of
34 kDa, and belongs to the few haemoglobins
which have a lower affinity for CO than for Oz.
Tracheal cell haemoglobin is also found in the
back-swimmers (Notonectidae) of the genera
Anisops and Buenoa. The haemoglobin of Anisops assimilis is a monomer of 16 kDa in the oxygenated state, and aggregates on deoxygenation.
The Hill plot is triphasic with n = 1.1, 5.2 and
2.6. The extremely high cooperativity in physiological conditions is explained entirely by the Ozdependent dissociation. In this case, the haemoglobin assists in the production of an Oz gas-bubble
for regulating buoyancy. Buenoa haemoglobin
also tends to aggregate. Intracellular haemoglobins have also been reported in the accessory
glands of the male genital tract of the water bug
Macrocorixa geoff roy and in the eggs of the louse
Pediculus humanus and the blood-sucking bug
Rhodnius prolixus [128, 174].
Extracellular haemoglobins are restricted to
the larvae of the non-biting midges (Chironomidae). These inhabit the mud regions of stagnant
water and appear red due to the haemoglobin dissolved in the haemolymph. As in the case of
Daphnia and Artemia, the haemoglobin concentration is higher in animals from low-Oz biotopes.
7.2.4 Haemoglobins of the Insects
273
The concentration in most species is highest during the fourth larval stage, and shows a marked
seasonal rhythm with a maximum in summer. In
extreme cases, haemoglobins make up more than
90 % of the haemolymph protein [104]. Contrary
to the rule that extracellular haemoglobins have
high molecular weights, the Chironomus haemoglobins are monomers or dimers, and seldom
tetramers (c. strenzkei) , of 16-kDa subunits.
Dimeric forms predominate in most species. All
the chironomid species possess 6-12 different
haemoglobins encoded by different genes. Twelve
haemoglobins have been described in the most
investigated species C. thummi thummi: these are
monomers CIT-I, -lA, -III, -IlIA, and -IV;
homodimers CTT-Il~, -VI, -VIlA, -VIlB, -VIII
and -IX; and CIT-X exists as monomers and
dimers in equilibrium [128].
The amino acid sequences are known for all
12 haemoglobins of C. thummi thummi [81, 108].
The chains vary in length between 136 and 151
amino acids due to extensions at both ends or to
deletions. Pairwise sequence comparisons give a
consistent 50 % agreement; 21 positions are
invariant if CIT-IlIA is not considered, but only
12 are conserved when this haemoglobin is
included in the analysis. Thus, all these haemoglobins apparently arose 200-400 million years
ago from a common monomeric ancestor. The
three-dimensional structure has been examined
only for CIT-III (Fig. 7.4) but, including all Hbridges, haem contacts etc., the analysis is at least
as thorough as for sperm whale myoglobin.
Dimer formation involves the interaction of G 19His (Lys in CTT-Il~) and G7-Glu. The monomeric forms contain G19-His but lack G7-Glu
[128]. The haem has the usual proximal contacts,
including F8-His, but the distal haem contacts are
unusual. With the exception of CIT-I, position
Ell is always occupied by isoleucine; this interacts with the haem iron and forces the E7-His out
of the haem pocket. CTT-I has valine at position
Ell and the Fe contact here is possibly via E7His. E7 in CIT-IlIA is occupied by glutamine.
The haemoglobins CIT-III and CTT-IV include
isomers in which the haem is rotated 180 0 , as has
already been described for vertebrate myoglobins
and the haemoglobin of the annelid Glycera
[31, 128].
It has been proposed that the site of haemoglobin biosynthesis in the chironomids lies not only
in the fat bodies but also in the oenocytes; however, tracer experiments have now shown that
haemoglobins are synthesized in subepithelial fat
bodies and temporarily stored in the oenocytes.