Are structural proteins in insect cuticles dominated by intrinsically disordered regions?

Fifty years ago it was concluded that the highly elastic cuticular protein, resilin, is devoid of secondary structure and that the peptide chains are randomly coiled and easily and reversibly deformed. These properties indicate that resilin is an intrinsically disordered protein and suggest that also other cuticular proteins may contain disordered regions. Amino acid sequences are now available for cuticular proteins from many insect species, and several programs have been developed to predict the probability for a given protein to contain disordered regions.

Studies on resilin-like gene products in insects.

Putative pro-resilins from 12 Drosophila species are compared with each other and with some pro-resilin-related proteins from other insect species, in an attempt to decide which structural features are likely to be important for the characteristic properties of resilins. The putative pro-resilins from the 12 Drosophila species are very similar; their structures are characterized by a chitin-binding R&R Consensus sequence of type RR-2, surrounded by two repeat-containing regions. The repeat-containing regions are assumed to be responsible for the long-range elasticity characteristic for resilin.

Insect cuticular sclerotization: a review.

Different regions of an insect cuticle have different mechanical properties, partly due to different degrees of stabilization and hardening occurring during the process of sclerotization, whereby phenolic material is incorporated into the cuticular proteins. Our understanding of the chemistry of cuticular sclerotization has increased considerably since Mark Pryor in 1940 suggested that enzymatically generated ortho-quinones react with free amino groups, thereby crosslinking the cuticular proteins. The results obtained since then have confirmed the essential features of Pryor's suggestion, and the many observations and experiments, which have been obtained, have led to a detailed and rather complex picture of the sclerotization process, as described in this review.

Quantitative determination of catecholic degradation products from insect sclerotized cuticles.

Acid hydrolysates of cuticle from various insect species were quantitatively analyzed for five catecholic amino acid adducts. Four of the adducts are ketocatechols; in three of them the amino acid moiety, either lysine, glycine or beta-alanine, is connected via its amino group to the alpha-carbon atom of 3,4-dihydroxyacetophenone, in the fourth a tyrosine residue is connected to the same position via its phenolic group. The fifth adduct contains histidine linked via its imidazole-ring to the beta-position of the dopamine sidechain.

Involvement of tyrosine residues, N-terminal amino acids, and beta-alanine in insect cuticular sclerotization.

During sclerotization of insect cuticle the acyldopamines, N-acetyldopamine (NADA) and N-beta-alanyldopamine (NBAD), are oxidatively incorporated into the cuticular matrix, thereby hardening and stabilizing the material by forming crosslinks between the proteins in the cuticular matrix and by forming polymers filling the intermolecular spaces in the cuticle. Sclerotized cuticle from the locust, Schistocerca gregaria, and the beetle, Tenebrio molitor, was hydrolyzed in dilute hydrochloric acid, and from the hydrolysates some components presumably degradation products of cuticular crosslinks were isolated. In two of the components, the sidechain of 3,4-dihydroxyacetophenone was linked to the amino groups of glycine and beta-alanine, respectively, and in the third component to the phenolic group of tyrosine.

Aspects of cuticular sclerotization in the locust, Scistocerca gregaria, and the beetle, Tenebrio molitor.

The number of reactive amino groups in cuticular proteins decreases during the early period of insect cuticular sclerotization, presumably due to reaction with oxidation products of N-acetyldopamine (NADA) and N-beta-alanyldopamine (NBAD). We have quantitated the decrease in cuticular N-terminal amino groups and lysine epsilon-amino groups during the first 24h of sclerotization in adult locusts, Schistocerca gregaria, and in larval and adult beetles, Tenebrio molitor, as well as the increase in beta-alanine amino groups in Tenebrio cuticle. The results indicate that nearly all glycine N-terminal groups and a significant part of the epsilon-amino groups from lysine residues are involved in the sclerotization process in both locusts and Tenebrio.

The extensible alloscutal cuticle of the tick, Ixodes ricinus.

The proteins in the distensible alloscutal cuticle of the blood-feeding tick, Ixodes ricinus, have been characterized by electrophoresis and chromatography, two of the proteins were purified and their total amino acid sequence determined. They show sequence similarity to cuticular proteins from the spider, Araneus diadematus, and the horseshoe crab, Limulus polyphemus, and to a lesser extent to insect cuticular proteins. They contain a conserved sequence region, which is closely related to the chitin-binding Rebers-Riddiford consensus sequence present in many insect cuticular proteins.

Chlorinated tyrosine derivatives in insect cuticle.

A method for quantitative measurement of 3-monochlorotyrosine and 3,5-dichlorotyrosine in insect cuticles is described, and it is used for determination of their distribution in various cuticular regions in nymphs and adults of the desert locust, Schistocerca gregaria. The two chlorinated tyrosine derivatives were present in all analyzed regions in mature adult locusts, the highest concentrations were found in the sclerotized cuticle of femur and tibia, but significant amounts were also present in the unsclerotized arthrodial membranes. Small amounts of the two amino acids were obtained from pharate, not-yet sclerotized cuticle of adult femur and tibia, the amounts increased rapidly during the first 24 h after ecdysis and more slowly during the next two weeks.

Regional differences in degree of resilin cross-linking in the desert locust, Schistocerca gregaria.

Various cuticular regions from the desert locust, Schistocerca gregaria, were quantitatively analyzed for two cross-linking amino acids, dityrosine and trityrosine, characteristic constituents of the rubberlike cuticular protein, resilin. These amino acids were found in all regions of cuticle investigated, but in widely varying amounts. In fully mature adult locusts the largest amounts of di- and trityrosine were obtained from the prealar arms and wing-hinges, structures possessing long-range elasticity and being involved in energy storage in the flight system.

Cuticular proteins from the horseshoe crab, Limulus polyphemus.

Proteins were purified from the carapace cuticle of a juvenile horseshoe crab, Limulus polyphemus, and several of them were characterized by amino acid sequence determination. The proteins are small (7-16 kDa) and their isoelectric points range from 6.5 to 9.

Sequence determination of three cuticular proteins and isoforms from the migratory locust, Locusta migratoria, using a combination of Edman degradation and mass spectrometric techniques.

The cuticle (exoskeleton) is a characteristic structure of insects and other arthropods. It is an extracellular layer which surrounds and protects the insect, and it is composed of proteins, lipids, water molecules, phenolic materials and chitin. Four proteins isolated from the thorax and femur cuticle of pharate adult migratory locust, Locusta migratoria, have been purified by ion-exchange chromatography and reversed-phase high performance liquid chromatography (RP-HPLC).