Properties and distribution of a chymotryptic-like enzyme from the sipunculan, Phascolopsis gouldii

Abstract

Homogenates of digestive tract, digestive fluid extracts, and a partially purified enzyme from Phascolopsis gouldii demonstrated activities toward N-benzoyl-L-tyrosine ethyl ester (BTEE) and N-glutaryl-L-phenylalanine p-nitroanilide (GPANA) at pH 7.8. The chymotryptic component showed optimal BTEE activity from pH 7.7 to 8.2 and was found to be stable at pH 5.0–9.0 with immediate and irreversible loss of activity at pH 4.0. BTEE activity was inhibited by L-1-tosylamide-2-phenylethyl chloromethyl ketone (TPCK), phenylmethylsulfonyl fluoride (PMSF), and phenothiazine-n-carbonyl chloride (PCC) and demonstrated affinity for 4-phenylbutylamine on a Sepharose 4B column. The molecular weight of the digestive fluid component as determined by gel chromatography was approximately 23 400. Gel chromatography of the eluant demonstrating pheynlbutylamine affinity yielded additional low molecular weight components. The posterior loop, which forms the junction between descending and ascending intestine, demonstrated highest BTEE and GPANA activities. In contrast, secretory cells are concentrated in the anterior descending intestine and granules appear to be emitted as an apocrine secretion.