Hydrolases in the mucosa of rat small intestine for phenylalanine-containing dipeptides

Abstract

A qualitative survey, made with a high-voltage paper electrophoresis screening technique, showed that mucosa of rat small intestine catalyzes the hydrolysis of 15 dipeptides containing 7 different amino acids. A recently described assay method was used for the quantitative determinations of hydrolase activities for Gly- l-Phe, Gly- l-Try, l-Met- l-Phe, l-Ala- l-Phe, l-Leu- l-Phe, Gly- l-Tyr, l-Phe- l-Met, l-Phe- l-Phe, l-Phe-Gly and l-phenylalanine amide in rat intestinal mucosa. The l-Ala- l-Phe hydrolase activity was studied extensively. It is rapidly inactivated above 45°, is unable to catalyze the hydrolysis of l-Ala l-Phe diketopiperazine and d-Ala- d-Phe, is only slightly active with d-Ala- l-Phe or l-Ala- d-Phe and is inhibited by p- chloromercuribenzoate but not by DFP or β-phenylpropionic acid. The soluble l-Ala- l-Phe hydrolase activity of intestinal mucosa requires a divalent cation, probably Zn 2+, for full activity and is distinct from the mucosal l-leucyl-β-naphthylamidase, Gly- l-Ala- l-Phe aminopeptidase, and l-Phe-Gly hydrolase activities. In mucosal homogenates approx. 90% of the activity of the l-Ala- l-Phe hydrolase, and of 6 of the other dipeptide hydrolases studied, is in the soluble fraction. The specific activity of soluble l-Ala- l-Phe hydrolase in intestinal mucosa is 30-fold higher than gastric, 4-fold greater than colonic and slightly less than renal specific activity. Specific activity in intestinal mucosa is 4-fold greater than in the remaining layers of the intestinal wall. Specific activities of hydrolases for l-Ala- l-Phe or l-Phe-Gly in mucosa from upper, middle and lower small intestine show no significant regional differences. l-Ala- l-Phe hydrolase activity of rat intestinal mucosa increases almost 4-fold between approximately the 7th and 25th days of life. Fasting a rat for 48 h does not significantly change the specific activity of l-Ala- l-Phe or l-Gly hydrolase of intestinal mucosa or kidney.