model do, in fact, exist, and the model is
accepted by a majority of researchers [4, 128,
138]. The alternative model explains sclerotization by non-polar interactions between protein
chains and an intervening melanin- or lignin-like
polymer. According to this model, the dehydration associated with sclerotization is the cause of
increased hydrophobic interactions between the
components and the higher mechanical strength
of the cuticula [4, 138]. The following arguments
are offered in favour of the second model: nuclear magnetic resonance (NMR) spectra of the
pupal cuticula of Manduca sexta have so far not
revealed any quinone bridges [182]. The number
of amino acids available for bridge building, i.e.
lysine residues and N-termini, amounts to only
one in 20-36 amino acids, and is insufficient to
account for the observed increase in mechanical
strength. Finally, it has been found that dehydration of isolated larval cuticulae from Calliphora
vomitoria always produces the same values for
tensile strength, irrespective of whether the cuticula is treated before or after sclerotization.
The continuing uncertainty about the basic
principles of sclerotization can be explained by
the extraordinary chemical and structural complexity of the insect cuticula and the multiplicity
of processes that occur. The individual processes
of sclerotization take place within nanoseconds to
several hours or days. Only part of the components involved are present before the process
begins; some appear only during sclerotization
and are very short-lived. Little is known about
the cuticular proteins involved in sclerotization
(p. 391), and it is also not at all clear how far quinones serve as bridges in the binding of protein
and chitin [128, 138].
Protein tanning by quinone was discovered in
1940 by Pryor in the egg cocoon of the cockroach
Blatta orientalis. In the cockroaches, the oviduct
ends in a genital cavity to which the paired colleterial glands are connected. The secretion of the
left gland contains dissolved proteins, diphenoloxidase and various catechol-j3-glucosides, and
that of the right gland contains a j3-glucosidase.
Depending upon the species, the catechol portion
of the glucosides consists of differing proportions
of 3,4-dihydroxybenzoic acid (protocatechuic
acid) and the corresponding aldehyde or alcohol
(Fig. 12.11). After cleavage of the glucoside bond,
the diphenol is oxidized to o-quinone, which causes the white solution of dissolved proteins to
solidify into the brown egg cocoon (ootheca). As
this contains no chitin, its mechanical strength
depends entirely upon the tanning of proteins.
12.6.2 The Process of Sc1erotization
431
The egg cocoon of the praying mantids is formed
in a similar manner. In this case, the left colleterial gland contains a j3-glucosidase, and the right
gland contains acyldopamine-j3-glucoside, with
species-specific differences in the acyl residues:
acetyl- in Statilia maculata, malonyl- and acetylin Tenodera aridifolia and T. angustipennis, and Nacetyl-l3-alanyl- in Mantis religiosa; Hierodula
petelliphera has these three plus N-I3-alanyl- and
N-malonyl-l3-alanyl- [276].
Considering the functional adaptations required, it is to be expected that sclerotization of
insect cuticulae is associated with great heterogeneity in the bridge structures between the polypeptides and between proteins and chitin fibrils
[138). The hydrolysis products of sclerotized cuticulae and in vitro experiments with catechols or
quinones, proteins and phenoloxidases give the
following picture of the process of bridge formation (Fig. 12.12a): N-acetyldopamine (NADA) is
oxidized to the corresponding quinone by phenoloxidase, and binds to the free amino group of a
protein to give a catechol-protein combination.
This is further oxidized to quinone and binds to a
second polypeptide to form a bridge, the structure of which is uncertain [4, 138]. Dopa- and
dopamine-quinone tend to form precursors of
melanin by intramolecular condensation; this
reaction is prevented in the sclerotizing cuticula
of insects by blocking of the amino group (Nacyldopamine) or its absence (protocatechic
acid). Quinone bridges can also be formed from
protein-bound dopa-quinone, which arises from a
tyrosine residue of the polypeptide chain by the
action of phenoloxidase (self-tanning). Finally,
cuticular proteins can also be linked by bistyrosine residues (see Fig. 11.4, p. 380) [138].
The formation of a completely different type of
bridge structure was described by Andersen in
1970, and this was termed p-sclerotization
because the protein in this case is bound to the 13
carbon atom of the catechol side-chain and not to
the ring. In the meantime, it has been found that
protein bonding can also occur via the a carbon
atom of the side-chain (Fig. 12.12b ), but the
name l3-sclerotization has been retained. Andersen assumed that the l3-sclerotizing agent was a,l3dehydro-NADA (Fig. 12.12b) , which was supposedly formed by a desaturase. On the other hand,
Sugumaran's research group favoured quinone
methide as the l3-sclerotizing agent and they
rechristened the process "quinone methide sclerotization". In fact, cuticular preparations from
various insects contain not only the quinone
methides of NADA and NBDA but also dehydro-
accepted by a majority of researchers [4, 128,
138]. The alternative model explains sclerotization by non-polar interactions between protein
chains and an intervening melanin- or lignin-like
polymer. According to this model, the dehydration associated with sclerotization is the cause of
increased hydrophobic interactions between the
components and the higher mechanical strength
of the cuticula [4, 138]. The following arguments
are offered in favour of the second model: nuclear magnetic resonance (NMR) spectra of the
pupal cuticula of Manduca sexta have so far not
revealed any quinone bridges [182]. The number
of amino acids available for bridge building, i.e.
lysine residues and N-termini, amounts to only
one in 20-36 amino acids, and is insufficient to
account for the observed increase in mechanical
strength. Finally, it has been found that dehydration of isolated larval cuticulae from Calliphora
vomitoria always produces the same values for
tensile strength, irrespective of whether the cuticula is treated before or after sclerotization.
The continuing uncertainty about the basic
principles of sclerotization can be explained by
the extraordinary chemical and structural complexity of the insect cuticula and the multiplicity
of processes that occur. The individual processes
of sclerotization take place within nanoseconds to
several hours or days. Only part of the components involved are present before the process
begins; some appear only during sclerotization
and are very short-lived. Little is known about
the cuticular proteins involved in sclerotization
(p. 391), and it is also not at all clear how far quinones serve as bridges in the binding of protein
and chitin [128, 138].
Protein tanning by quinone was discovered in
1940 by Pryor in the egg cocoon of the cockroach
Blatta orientalis. In the cockroaches, the oviduct
ends in a genital cavity to which the paired colleterial glands are connected. The secretion of the
left gland contains dissolved proteins, diphenoloxidase and various catechol-j3-glucosides, and
that of the right gland contains a j3-glucosidase.
Depending upon the species, the catechol portion
of the glucosides consists of differing proportions
of 3,4-dihydroxybenzoic acid (protocatechuic
acid) and the corresponding aldehyde or alcohol
(Fig. 12.11). After cleavage of the glucoside bond,
the diphenol is oxidized to o-quinone, which causes the white solution of dissolved proteins to
solidify into the brown egg cocoon (ootheca). As
this contains no chitin, its mechanical strength
depends entirely upon the tanning of proteins.
12.6.2 The Process of Sc1erotization
431
The egg cocoon of the praying mantids is formed
in a similar manner. In this case, the left colleterial gland contains a j3-glucosidase, and the right
gland contains acyldopamine-j3-glucoside, with
species-specific differences in the acyl residues:
acetyl- in Statilia maculata, malonyl- and acetylin Tenodera aridifolia and T. angustipennis, and Nacetyl-l3-alanyl- in Mantis religiosa; Hierodula
petelliphera has these three plus N-I3-alanyl- and
N-malonyl-l3-alanyl- [276].
Considering the functional adaptations required, it is to be expected that sclerotization of
insect cuticulae is associated with great heterogeneity in the bridge structures between the polypeptides and between proteins and chitin fibrils
[138). The hydrolysis products of sclerotized cuticulae and in vitro experiments with catechols or
quinones, proteins and phenoloxidases give the
following picture of the process of bridge formation (Fig. 12.12a): N-acetyldopamine (NADA) is
oxidized to the corresponding quinone by phenoloxidase, and binds to the free amino group of a
protein to give a catechol-protein combination.
This is further oxidized to quinone and binds to a
second polypeptide to form a bridge, the structure of which is uncertain [4, 138]. Dopa- and
dopamine-quinone tend to form precursors of
melanin by intramolecular condensation; this
reaction is prevented in the sclerotizing cuticula
of insects by blocking of the amino group (Nacyldopamine) or its absence (protocatechic
acid). Quinone bridges can also be formed from
protein-bound dopa-quinone, which arises from a
tyrosine residue of the polypeptide chain by the
action of phenoloxidase (self-tanning). Finally,
cuticular proteins can also be linked by bistyrosine residues (see Fig. 11.4, p. 380) [138].
The formation of a completely different type of
bridge structure was described by Andersen in
1970, and this was termed p-sclerotization
because the protein in this case is bound to the 13
carbon atom of the catechol side-chain and not to
the ring. In the meantime, it has been found that
protein bonding can also occur via the a carbon
atom of the side-chain (Fig. 12.12b ), but the
name l3-sclerotization has been retained. Andersen assumed that the l3-sclerotizing agent was a,l3dehydro-NADA (Fig. 12.12b) , which was supposedly formed by a desaturase. On the other hand,
Sugumaran's research group favoured quinone
methide as the l3-sclerotizing agent and they
rechristened the process "quinone methide sclerotization". In fact, cuticular preparations from
various insects contain not only the quinone
methides of NADA and NBDA but also dehydro-