title: Intestinal surface peptide hydrolases: identification and characterization of three enzymes from rat brush border.
abstract: 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.
mesh_terms: D000626:Aminopeptidases / Q000378:metabolism; D000818:Animals; D002021:Buffers; D002413:Cations, Divalent; D004492:Edetic Acid / Q000494:pharmacology; D005260:Female; D005998:Glycine / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000201:enzymology*; D007421:Intestine, Small / Q000201:enzymology*; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D013329:Structure-Activity Relationship
chemical_list: D002021:Buffers; D002413:Cations, Divalent; D004492:Edetic Acid; D010447:Peptide Hydrolases; D000626:Aminopeptidases; D005998:Glycine