Hopkins 9418129 Insect cuticle sclerotization is a vital process that occurs periodically during each stage of insect development to harden and stabilize the newly secreted cuticular exoskeleton. The structural polymers of protein and chitin that make up the bulk of the cuticle, and their chemical interactions with quinone tanning agents are largely responsible for the final properties of the exoskeleton. Sclerotization is thought to involve the formation of covalent cross-links between protein nucleophilic side chain groups and the ring or side-chain carbons of N-acetyldopamine and N-Beta- alanyldopamine. Purified cuticular proteins from the pupal cuticle of the tobacco hornworm, Manduca sexta, involved in sclerotization reactions and their post-translational modification by quinonoid metabolites will be characterized as to molecular weights, amino acid compositions and sequences, carbohydrate components and catechol-amino acid adducts. In vitro tanning reactions will be studied between N-acyldopamine quinonoid derivatives and model compounds that have nucleophilic functional groups representative of amino acid residues in cuticular proteins. The adducts formed will be chemically characterized by electrochemistry, HPLC, NMR and mass spectrometry. Solid-state NMR studies utilizing 13C-, 15N-, 17O labeled protein and catecholamine precursors of cuticle, which will provide information on the nature of the covalent bonds formed in situ between the structural components during sclerotization. %%% Insect cuticle sclerotization is a vital process that occurs periodically during insect development to harden and stabilize the newly secreted cuticular exoskeleton. The exoskeleton serves many functions including protection, locomotion , respiration and communication, and therefore must have very diverse mechanical and chemical properties to provide for each function. The structural polymers of protein and chitin that make up the bulk of the cuticle, and their che mical interactions with quinone tanning agents derived from catecholamines are largely responsible for the final properties of the exoskeleton. Purified cuticular proteins from the pupal cuticle of the tobacco hornworm, Manduca sexta, involved in sclerotization reactions will be characterized and their modification by quinonoid metabolites determined. The products formed will be identified as to the types of chemical bonding involved in sclerotization. Solid-state NMR studies will provide a non-destructive method for investigating the molecular assembly of the exoskeleton. Understanding how the multifunctional insect exoskeleton is sclerotized at each stage of development is of major importance to insect biology and may facilitate the development is of new approaches to selective insect pest control. ***
