Separation and identification of cathepsins in newborn rat epidermis

Cathepsins B, D, H, and L were identified in the extract of 2-day-old rat epidermis and separated by gel filtration from aminoendopeptidase with a Mr of 400,000 and from the low-molecular-weight cysteine proteinase inhibitor. They were further purified by ion exchange column chromatography. The final separation for cathepsins B and H was performed by gel filtration, while cathepsin D was purified by pepstatin affinity chromatography and cathepsin L by fast protein liquid chromatography (FPLC). Substrate specificity, inhibitor susceptibility, and apparent molecular weights of the separated proteinases were determined and values compared to rat liver enzymes. Apparent molecular weights for epidermal cathepsins B, H, and L were higher than those for comparable liver enzymes of adult rats. The cysteine proteinase inhibitor in epidermis was found to inhibit cathepsins B, H, and L but not cathepsin D and aminoendopeptidase of rat epidermis. This study demonstrates the presence of cathepsin L in the epidermis and describes simultaneous separation and comparison of epidermal catheptic proteinases.