192
5 Plasma Proteins, Yolk Proteins and Metal-Binding Proteins
ent hexamers. Only one type of arylphorin monomer is found in the lepidopterans Calpodes ethlius, Heliothis zea, Hyalophora cecropia and Papilio polyxenes and in the honey bee Apis melli/era;
two occur in the lepidopterans Bombyx mori,
Galleria melonella and Manduca sexta and in the
cockroach Blatta orientalis; and three occur in the
dipterans Drosophila melanogaster, Ceratitis capitata and Musca domestica. In Sarcophaga peregrina, Lucilia styga and Calliphora vicina there are
several other components in addition to the main
subunit. The genes for the LHP subunits form a
mUlti-gene family that in the fly Calliphora
vicina, for example, includes 20 members. The
heterogeneity of the LHPs is increased even further by genetic variability, in some cases to a quite
unusual degree. Thus, no less than 90 LHP phenotypes were found in 180 flies of the species
Lucilia cyprina from an Australian population,
the genetic analysis of which indicated at least 12
homologous genes. The random combination of
isoforms and allelic variants should result in a
host of different LHPs [121]. LHP cDNAs or
genes from the dipterans S. peregrina and D. melanogaster and from the lepidopterans B. mori,
M. sexta and Heliothis virescens have been completely or partially sequenced [79, 121, 140, 187,
272]. The two LHP subunits a and ~ from
M. sexta agree by 68 % in their 686-687 amino
acids. Surprisingly, the Manduca LHP is significantly homologous to the respiratory blood pigment haemocyanin from the crab Panulirus interruptus. The highest sequence similarity is found
in the region of the polypeptide chain which, in
both proteins, forms the contact between the subunits of the hexamers [272]. Stage-specific
expression of arylphorin genes is regulated in the
fly Sarcophaga peregrina by a protein which only
appears in 46-hour-old larvae and binds specifically to the sequence ACCACAACA in position
-255 to -247 [130].
Apart from the arylphorins, a second type of
larval haemolymph protein is found in the lepidopterans; these contain 4-8 % methionine and
are therefore known as "methionine-rich storage
proteins". They are synthesized only in the final
larval stage and reach higher concentrations in
female than in male animals. Spodoptera litura
has two proteins of this type, whereas Bombyx
mori and Hyalophora cecropia have only one.
The methionine-rich proteins are significantly
homologous to the arylphorins and have the same
hexameric structure [121]. In B. mori the
methionine-rich protein SP-l agrees in 30 % of its
sequence with the arylphorin SP-2 [79]. Drosophila melanogaster possesses a larval haemolymph protein, LSP-2, which is rich in aromatic
amino acids but shows no immunological crossreactivity to the arylphorins. Animals with null
alleles for LSP-2 mostly die during development,
and in any case remain sterile. Similar proteins
are found in other dipterans [121]. In addition to
the typical arylphorin and methionine-rich proteins, there is a further haemolymph protein, found
in the pupae of Hyalophora cecropia, which is
rich in histidine and contains bound riboflavin
and copper. A further hexameric haemolymph
protein of ca. 500 kDa, which is present in the larvae of the migratory locust Locusta migratoria
but is missing in the adult, fits into none of the
known classes of haemolymph proteins in holometabolic insects. There are, in all probability,
further classes of as yet unknown larval haemolymph proteins in other insect groups [121].
5.6 Plasma Lipoproteins
In order to be transported between the sites of
resorption, de novo synthesis, conversion, and
use, hydrophobic lipids must be solubilized by
binding to proteins of the blood plasma. Consequently, transport lipoproteins are to be expected
in many animals but have so far only been examined in any detail in vertebrates and insects.
Both groups of animals contain spherical lipidprotein particles which can, in each case, be
resolved by electron microscopy but which have
very different molecular architectures. In vertebrates, there is a nucleus of non-polar lipids surrounded by a coat of proteins and polar lipids
(phospholipids and cholesterol); in insects, on
the other hand, the nucleus of the lipoprotein particles consists mainly of proteins and is encased
predominantly by relatively polar lipids (diacylglycerol). Specific lipoproteins, viteUogenins, are
found in the blood plasma of animals that produce yolked eggs, and are also incorporated into
the yolk material in more or less modified forms.
Three methods are usually used to examine lipoproteins: electrophoresis and lipid-specific staining give information about the availability and
variety of the lipoproteins, and density gradient
centrifugation or fractionated flotation in media
of differing density by the help of ultracentrifugation serves for their isolation. This takes advantage of the reduction in density of particles which
occurs with increasing lipid content. In the investigation of human lipoproteins, the density
5 Plasma Proteins, Yolk Proteins and Metal-Binding Proteins
ent hexamers. Only one type of arylphorin monomer is found in the lepidopterans Calpodes ethlius, Heliothis zea, Hyalophora cecropia and Papilio polyxenes and in the honey bee Apis melli/era;
two occur in the lepidopterans Bombyx mori,
Galleria melonella and Manduca sexta and in the
cockroach Blatta orientalis; and three occur in the
dipterans Drosophila melanogaster, Ceratitis capitata and Musca domestica. In Sarcophaga peregrina, Lucilia styga and Calliphora vicina there are
several other components in addition to the main
subunit. The genes for the LHP subunits form a
mUlti-gene family that in the fly Calliphora
vicina, for example, includes 20 members. The
heterogeneity of the LHPs is increased even further by genetic variability, in some cases to a quite
unusual degree. Thus, no less than 90 LHP phenotypes were found in 180 flies of the species
Lucilia cyprina from an Australian population,
the genetic analysis of which indicated at least 12
homologous genes. The random combination of
isoforms and allelic variants should result in a
host of different LHPs [121]. LHP cDNAs or
genes from the dipterans S. peregrina and D. melanogaster and from the lepidopterans B. mori,
M. sexta and Heliothis virescens have been completely or partially sequenced [79, 121, 140, 187,
272]. The two LHP subunits a and ~ from
M. sexta agree by 68 % in their 686-687 amino
acids. Surprisingly, the Manduca LHP is significantly homologous to the respiratory blood pigment haemocyanin from the crab Panulirus interruptus. The highest sequence similarity is found
in the region of the polypeptide chain which, in
both proteins, forms the contact between the subunits of the hexamers [272]. Stage-specific
expression of arylphorin genes is regulated in the
fly Sarcophaga peregrina by a protein which only
appears in 46-hour-old larvae and binds specifically to the sequence ACCACAACA in position
-255 to -247 [130].
Apart from the arylphorins, a second type of
larval haemolymph protein is found in the lepidopterans; these contain 4-8 % methionine and
are therefore known as "methionine-rich storage
proteins". They are synthesized only in the final
larval stage and reach higher concentrations in
female than in male animals. Spodoptera litura
has two proteins of this type, whereas Bombyx
mori and Hyalophora cecropia have only one.
The methionine-rich proteins are significantly
homologous to the arylphorins and have the same
hexameric structure [121]. In B. mori the
methionine-rich protein SP-l agrees in 30 % of its
sequence with the arylphorin SP-2 [79]. Drosophila melanogaster possesses a larval haemolymph protein, LSP-2, which is rich in aromatic
amino acids but shows no immunological crossreactivity to the arylphorins. Animals with null
alleles for LSP-2 mostly die during development,
and in any case remain sterile. Similar proteins
are found in other dipterans [121]. In addition to
the typical arylphorin and methionine-rich proteins, there is a further haemolymph protein, found
in the pupae of Hyalophora cecropia, which is
rich in histidine and contains bound riboflavin
and copper. A further hexameric haemolymph
protein of ca. 500 kDa, which is present in the larvae of the migratory locust Locusta migratoria
but is missing in the adult, fits into none of the
known classes of haemolymph proteins in holometabolic insects. There are, in all probability,
further classes of as yet unknown larval haemolymph proteins in other insect groups [121].
5.6 Plasma Lipoproteins
In order to be transported between the sites of
resorption, de novo synthesis, conversion, and
use, hydrophobic lipids must be solubilized by
binding to proteins of the blood plasma. Consequently, transport lipoproteins are to be expected
in many animals but have so far only been examined in any detail in vertebrates and insects.
Both groups of animals contain spherical lipidprotein particles which can, in each case, be
resolved by electron microscopy but which have
very different molecular architectures. In vertebrates, there is a nucleus of non-polar lipids surrounded by a coat of proteins and polar lipids
(phospholipids and cholesterol); in insects, on
the other hand, the nucleus of the lipoprotein particles consists mainly of proteins and is encased
predominantly by relatively polar lipids (diacylglycerol). Specific lipoproteins, viteUogenins, are
found in the blood plasma of animals that produce yolked eggs, and are also incorporated into
the yolk material in more or less modified forms.
Three methods are usually used to examine lipoproteins: electrophoresis and lipid-specific staining give information about the availability and
variety of the lipoproteins, and density gradient
centrifugation or fractionated flotation in media
of differing density by the help of ultracentrifugation serves for their isolation. This takes advantage of the reduction in density of particles which
occurs with increasing lipid content. In the investigation of human lipoproteins, the density