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The dipeptidase activity toward L-glutaminyl-L-proline and glycyl-L-proline has been studied in the small intestinal mucosa of human fetuses and newborns between 14 and 36 weeks of fetal age.

Extract: The dipeptidase activity toward L-glutaminyl-L-proline and glycyl-L-proline has been studied in the small intestinal mucosa of human fetuses and newborns between 14 and 36 weeks of fetal age.The small intestine was removed within 15 minutes of death. It was immediately frozen and used for enzymatic studies within 2 to 30 days. The activity of both enzymes was stable for as long a period as three months at a temperature of -20°. Mucosal scrapings were homogenized in 0.01 M Tris-HCl buffer, pH 7, and then centrifuged at 1500 × g for 10 minutes. The supernatant, which contained more than 95% of the enzyme activity, was used for assay; it was incubated in 20 mM substrate, at optimum pH, under conditions permitting calculation of initial velocity. Free proline was measured at the end of the incubation period. By the 14th to 16th week of age, levels of activity of both enzymes were comparable with those of the adult (fig. 1).In order to investigate the distribution of dipeptidase activity, the small bowel from fetuses between 22 and 34 weeks of age was examined. Each sample was homogenized separately before studying. Uniformly high levels of dipeptidase activity were found throughout the proximal six-tenths of the small bowel, while lower values were observed in the terminal ileum (fig. 2). A significant difference in the activity of both enzymes was found between the proximal and the distal third of the small bowel when calculated according to Student's t-test, p < 0.02 (table I). The activity ratios were constant throughout the length of the intestine during the entire age span studied.After centrifugation of the homogenate of intestine at 105,000 × g for one hour, gel filtration on Sephadex G 200 of the supernatant resulted in the partial purification of the enzyme, as indicated by a 4.5- to 6.5-fold increase in specific activity.The enzymes of the crude and the partially purified extracts were studied to determine the influence on enzyme activity of pH, Co++and Mn++ions, heat treatment, substrate concentrations. Co++and Mn++ions clearly activated hydrolysis of glycyl-L-proline, but did not affect that of L-glutaminyl-L-proline. Heating at 40° in a solution of 0.002 M Mn++resulted in an increase of activity of glycyl-L-proline dipeptidase and the nearly complete disappearance of that of L-glutaminyl-L-proline dipeptidase (fig. 5); however, the activity of both enzymes was not separated by gel filtration, ammonium sulphate precipitation, or DEAE cellulose chromatography. Further studies are necessary to determine if a specific enzyme (s) of the small bowel mucosa, other than glycyl-L-proline dipeptidase (EC.3.4.3.7), causes hydrolysis of L-glutaminyl-L-proline.Glycyl-L-proline dipeptidase activity did not change during the age period studied; however, L-glutaminyl-L-proline dipeptidase activity of 14- to 24-week-old fetuses displayed a pH dependence and a kinetic versus substrate concentration not observed in fetuses from 26 to 36 weeks of age (fig.4). These results may suggest that a qualitative change of L-glutaminyl-L-proline dipeptidase had occurred with maturation.Speculation: This study suggests that in the small intestine of the human fetus, L-glutaminyl-L-proline dipeptidase in early stages of life differs qualitatively from that of older fetuses. Further studies on pure enzymes may elucidate the basic mechanism that underlies qualitative changes of some enzyme molecules during growth.