Assembly of nematode cuticle: role of hydrophobic interactions in CUT-2 cross-linking.

CUT-2 is a component of cuticlin, the highly cross-linked, insoluble residue of the cuticle of the nematode Caenorhabditis elegans. A recombinant fragment of CUT-2, produced in E. coli, can be cross-linked in vitro by horse radish peroxidase via dityrosine formation to give large molecular species [1]. In this paper it is shown that the formation of CUT-2 polymers is greatly favoured over that of CUT-2 oligomers as no low molecular weight intermediates, dimers or trimers can be detected even when the cross-linking reaction is slowed or interrupted before completion. This suggests that recombinant CUT-2 forms large non-covalent complexes that are the only competent substrate for cross-linking. The inhibition of cross-linking by urea and the behavior of recombinant CUT-2 in size-exclusion chromatography under a variety of conditions suggest that hydrophobic interactions are important in the formation and stabilization of these complexes. The complexes are excellent substrates for cross-linking but react poorly with free tyrosine. In contrast, a soluble recombinant CUT-2 is a poor substrate for cross-linking but can efficiently react with free tyrosine.