Structural proteins in the mammalian epidermis contain citrulline residues generated by enzymatic deimination of arginine residues. We analyzed deiminated proteins solubilized from sequentially stripped layers of guinea pig epidermis. Deiminated proteins were localized in the granular and cornified layers. Those in the inner layer enriched with granular cells were resolved into numerous components by two-dimensional gel electrophoresis. An arc-shaped high-molecular-weight smear and two series of charged isomers among them coincided with filaggrin immunoreactivity. Several groups of filaggrin-negative spots appeared to be generated by further deimination and proteolysis of these filaggrins. Deiminated protein spots co-migrating with type II and type I keratins were also detected. Deiminated filaggrins and their further processed derivatives disappeared in the outer layer, while deiminated keratins persisted. These data suggested that filaggrin as well as profilaggrin were deiminated during the posttranslational processing in guinea pig skin, and that some keratins were deiminated preferentially during the cornification of epidermis, Possible biological significance of protein deimination in guinea pig skin was discussed in comparison with our recent finding on deiminated proteins in rat skin.