Properties of a trypsin and chymotrypsin inhibitor secreted by larval Taenia pisiformis

Living metacestodes of Taenia pisiformis maintained in vitro discharge into the surrounding medium a protease inhibitor, which has been purified from the medium by affinity chromatography on bovine α-chymotrypsin immobilized to CNBr-activated Sepharose 4B. The purified inhibitor was shown to inactivate the hydrolysis of $\text{N-α-}\text{benzoyl}\text{-}\text{L}\text{-}\text{arginine}$ ethyl ester and $\text{N-}\text{benzoyl}\text{-}\text{L}\text{-}\text{tyrosine}$ ethyl ester, respectively, by trypsin and chymotrypsin of bovine, rabbit and dog origin, and also the hydrolysis of casein by both bovine trypsin and bovine α and β chymotrypsins, but it did not affect the enzymic activity of subtilisin, elastase, collagenase, pepsin, rennin and papain. The inhibitor withstood heating at 100°C for up to 30 min, was stable in the pH range of 1.5–8.0, was unaffected by 8 M-urea or 0.2 M-2-mercaptoethanol, and had a molecular weight of about 7000 as calculated from its gel chromatographic behaviour. The inhibitor specifically inhibits either trypsin or chymotrypsin with the formation of stable enzyme inhibitor complexes that are not dissociated by 4 M-KCl. Inhibition of trypsin and chymotrypsin is non-competitive and is linear with inhibitor concentration up to 70–80% inhibition. Inhibitory activities toward both enzymes are functions of the same binding site of the inhibitor molecule. Complex formation between the inhibitor and the enzymes is timedependent; it requires 3–4 min for completion.