| Summary: | Peptide hydrolases were solubilized from rat small intestinal brush border by papain and separated by Sephadex G-200 chromatography, velocity gradient ultracentrifugation and polyacrylamide disc electrophoresis and designated according to approximate molecular size from sedimentation studies. Peptidases I (apparent Mr 230 000) and II (apparent Mr 160 000) are oligopeptidases with maximum specificity for tripeptides with identical pH optima (7.5) and similar apparent Km with L-Leu-Gly (I, 0.60 MM; II, 0.76 mM). L-Leucyl-beta-naphthylamide is a competitive inhibitor of both enzymes. Concentration of peptidase II produced partial conversion to peptidase I on polyacrylamide disc electrophoresis. The third peptide hydrolase (III, Mr 120 000) is a dipeptidase with pH optimum 8.5 and apparent Km for L-Leu-Gly of 0.65 mM. These peptide hydrolases were inhibited appreciably (37-59%) by 0.2 M glycine/NaOH, Tris - HCl or Tris - glycine buffers. EDTA (5 mM) completely inhibited these enzymes but all activity was restored by dialysis against buffer without divalent ions. Subsequent addition of Mg2+, Mn2+, Co2+ or Zn2+ (1-2 mM) inhibited peptidases I and II variably (4-81%) depending upon the substrate and buffer used. In contrast peptidase III was activated slightly by metal ions (5-20%). These peptide hydrolases are strategically located at the intestinal lumen-cell interface and possess biochemical characteristics making them ideally suited to play a pivotal role in the final stage of protein digestion.
|
|---|