IDENTIFICATION AND CHARACTERIZATION OF A PRO-TUMOR NECROSIS FACTOR-ALPHA-PROCESSING ENZYME FROM THE ADAM FAMILY OF ZINC METALLOPROTEASES

Tumor necrosis factor-alpha (TNF) is initially expressed as a 26-kDa m embrane-bound precusor protein (pro-TNF) that is shed proteolytically from the cell surface, releasing soluble 17-kDa TNF. We have identifie d human ADAM 10 (HuAD10) from THP-1 membrane extracts as a metalloprot ease that specifically clips a peptide substrate spanning the authenti c cleavage site between Ala(76) and Val(77) in pro-TNF. To confirm tha t HuAD10 has TNF processing activity, we cloned, expressed, and purifi ed an active, truncated form of HuAD10. Characterization of recombinan t HuAD10 (rHuAD10) suggests that this enzyme has many of the propertie s (i.e. , substrate specificity, metalloprotease activity, cellular lo cation) expected for a physiologically relevant TNF-processing enzyme.