Dipeptidyl Arylamidase III of the Pituitary

Abstract

Abstract An aminopeptidase has been demonstrated in the bovine anterior pituitary which hydrolyzes dipeptidyl residues from the amino terminus of peptides containing a minimum of 4 residues. Tetraalanine and hexaalanine are completely hydrolyzed to dialanine, whereas pentaalanine is cleaved to dialanine and trialanine. N-Acetyltetraalanine is not hydrolyzed. Similarly, tetralysine and tetraphenylalanine are hydrolyzed to the respective dipeptides; tetraglutamic acid and tetraglycine are not attacked. Of the dipeptidyl-β-naphthylamides (βNA), only Arg-Arg-βNA is readily hydrolyzed, yielding Arg-Arg and β-naphthylamine, and appears to serve as a relatively specific substrate since Ala-Ala-βNA, Lys-Lys-βNA, Gly-Arg-βNA, Ser-Tyr-βNA, Leu-Ala-βNA, and Lys-Ala-βNA are not attacked. The optimum for the hydrolysis of the susceptible substrates lies at pH 8.5 to 9.0. The maximal rate of hydrolysis of Arg-Arg-βNA occurs at 0.04 mm; greater concentrations of substrate as well as the product, Arg-Arg, are inhibitory. The enzyme is inhibited by thiol reagents and activated by thiol compounds. A 500-fold purification can be achieved by fractionation with (NH4)2SO4 and chromatography on diethylaminoethyl cellulose.