2 DISTINCT FORMS OF HUMAN MAST-CELL CHYMASE - DIFFERENCES IN AFFINITYFOR HEPARIN AND IN DISTRIBUTION IN SKIN, HEART, AND OTHER TISSUES

Chymase, a chymotrypsin-like protease secreted by human mast cells, is generally considered to be a single enzyme. However, by heparin-agaro se chromatography of high-salt extracts of human skin, we have consist ently resolved three peaks of chymotryptic activity, eluting at 0.4 M NaCl (peak A), 1.0-1.2 M NaCl (peak B) and 1.8-2.0 M NaCl (peak C), wi th peak B containing 75-90 % of the recovered activity. Each peak reta ined its identity upon rechromatography. The three peaks of activity w ere similar in substrate specificity and inhibitor profile and distinc tly different from other chymotryptic enzymes, including cathepsin G a nd the stratum corneum chymotryptic enzyme. Examination of different t issues revealed that peak C was virtually absent from synovial tissue, was present as a minor component in skin and heart, but constituted t he predominant chymotryptic activity in lung. Peaks B and C from skin tissue were further purified by chromatography on Sephacryl S-200. Bot h had a molecular mass of 28-29 kDa, yielded the N-terminal sequence r eported for chymase, and on western blots reacted with a panel of poly clonal, monoclonal and antipeptide antibodies against chymase. Chymase C required higher concentrations of NaCl to overcome the stimulatory effects of heparin than did chymase B, but had a similar pH profile. T hus, human chymase exists in at least two distinct but similar forms, and the differences in heparin binding and tissue distribution could h ave important consequences for enzyme function.