(13f) Enzymatic Reactions and Mechanical Properties of Beetle Elytral Cuticle, a Multicomponent Biomaterial

Fully tanned insect cuticle or exoskeleton is a strong, lightweight biomaterial comprised primarily of chitin, proteins and catechols. It has been estimated that the chitin fibers embedded within the cuticle have a predicted modulus of 250 GPa, comparable to metal alloys and high performance carbon fiber composites. To design a strong biocompatible material based on these superb natural properties, the origin of the high strength of cuticle must be understood.

Dynamic mechanical analysis was used to evaluate the mechanical properties of elytral (wing cover) cuticle of the Tribolium castaneum (red flour beetle) and to examine the relationships between the different components of this complex biomaterial. This examination of cuticle mechanical performance as a function of multiple variables aims to characterize the potential of this biomaterial and to shed light on the roles and interactions of the primary cuticle components chitin, protein, catechol and water. By means of comprehensive mechanical characterization, the importance upon cuticle maturation of cross-link formation relative to loss of plasticization by water due to dehydration is assessed, suggesting the utility of such analysis in evaluation of microscopic organization in biological materials.

Enzymatic pathways involved in the maturation of elytral cuticle of Tribolium castaneum have been investigated by altering enzymatic activity using RNA interference and correlating pathway hypotheses with quantitative evaluation of a comprehensive set of elytral mechanical properties. Use of RNAi to suppress the expression of two different enzymes involved in tanning, laccase2 (Lac2) and aspartate 1-decarboxylase (ADC), led to very different changes in cuticle. Lac2 knockdown produced weak, highly hydrated but elastic cuticle, while ADC knockdown induced formation of a strong but more pigmented cuticle with a larger viscous component to its dynamic mechanical properties. These results are consistent with the hypothesis that ADC inhibition favors formation of melanic pigment together with a decrease in protein cross-linking, whereas Lac2 suppression inhibits both processes. The combination of biochemical techniques with materials characterization methods is a powerful means of evaluating the roles of different enzymes in cuticle development.