PURIFICATION AND CHARACTERIZATION OF LEUKOTRIENE A(4) HYDROLASE FROM HUMAN EPIDERMIS

The leukotriene A(4) hydrolase is a central enzyme in leukotriene B-4 formation. Unlike 5-lipoxygenase, leukotriene A(4) hydrolase activity is present in normal human epidermis, where it is likely to be involve d in transcellular leukotriene formation. In this study the leukotrien e A(4) hydrolase was purified from human epidermis and human cultured keratinocytes and compared with leukotriene A(4) hydrolase from human neutrophils. To purify leukotriene A(4) hydrolase from human epidermis a new non-specific affinity chromatography column, with the leukotrie ne A(4) hydrolase inhibitor bestatin coupled to AH-Sepharose, was intr oduced. The epidermal leukotriene A(4) hydrolase was purified to appar ent homogeneity and the molecular weight was determined to be approxim ately 70,000 Da by SDS-PAGE. The pI was 5.1-5.4 for the epidermal as w ell as the keratinocyte and neutrophil leukotriene A(4) hydrolase, as determined by chromatofocusing. Only minor differences in the amino ac id composition were seen between the three enzyme sources. The optimal pH for the hydrolase activity was 7.5-8.5 for the epidermal and neutr ophil leukotriene A(4) hydrolases. Finally, it was also shown that the epidermal leukotriene A(4) hydrolase undergoes suicide inactivation w hen transforming leukotriene A(4) into leukotriene B-4. It was conclud ed that there is a close resemblance between the epidermal leukotriene A(4) hydrolase and the hydrolase found in other cell types. Therefore , the human epidermis may be a good model for the in vivo study of tra nscellular leukotriene formation.