Transglutaminase reactivity of human involucrin.

Human involucrin (hINV) is assembled into cornified structures via formation of transglutaminase (TG)-dependent interprotein epsilon-(gamma-glutamyl)lysine bonds. The hINV sequence includes 150 glutamine residues that could function as potential sites of cross-link formation. The present studies were designed to evaluate the extent to which hINV can function as a TG substrate under optimal conditions and in the absence of other substrates. Incubation of hINV with TG results in formation of 4-5 isopeptide bonds per hINV molecule. When the small amine donor (14)C-putrescine is included in the reaction, 48 Q residues are labled. Isotope distribution and sequence analysis suggests that the (14)C-putrescine-labeled sites are located throughout the protein. Our present results show that many hINV Q residues can be utilized for cross-link formation, and that hINV can be cross-linked at very high cross-link densities. These results suggest that, in vivo, factors other than hINV structure limit the number of residues used for cross-link formation.