Keratinization is an orderly process in which protein-containing monomers are synthesized and assembled in viable epidermal cells, keratinocytes. The final product, keratin, is not secreted but is retained within the cellular envelope in non-viable aneucleate cells at the skin surface. Recently, covalent cross-links were identified in wool keratin and shown to be epsilon-(gamma-glutamyl) lysine bonds. Transglutaminase, an enzyme which catalyzes the formation of gamma-glutamyl-epsilon-lysine bonds, has been isolated in soluble form and purified from guinea pig hair follicles and bovine and rat epidermis. We propose to analyze epidermal transglutaminase (ET) in the skin of experimental animals (cow and rat) and man. ET will be purified by chromatographic and electrophoretic methods and assayed by its ability to incorporate the fluorescent substrate, dansyl cadaverine, into casein. Keratinocyte transglutaminase will be strictly compared to and distinguished from plasma transglutaminase (fibrin stabilizing factor, factor XIII) by enzymatic, physical and immunochemical properties. The enzyme ET will be localized in epidermis and epidermal appendages by histochemical or immunochemical methods by light- or electron-microscopy. ET will be employed as a biochemical probe to identify native soluble donor and acceptor proteins in epidermis.
