The presence of insulin-degrading enzyme in human ileal and colonic mucosal cells.
The aim of this research is to characterize the presence of insulin-degrading enzyme in human colon and ileal mucosal cells. Biochemical studies, including the activity-pH profiles, the effects of enzyme inhibitors, immunoprecipitation and western blots, were conducted. The majority of insulin-degrading activity in colon mucosal cells was localized in the cytosol. In both colon and ileum, cytosolic insulin-degrading activities had a pH optimum at pH 7.5, and were extensively inhibited by each of N-ethylmaleimide, p-chloromercuribenzoate, and 1,10-phenanthroline, but were very weakly affected by each of leupeptin, chymostatin, diisopropyl phosphofluoridate and soybean trypsin inhibitor. In the colon and ileum, more than 93% and 96%, respectively, of cytosolic insulin-degrading activities were removed by the mouse monoclonal antibody to human RBC insulin-degrading enzyme, as compared with less than 20% by the normal mouse IgG for both tissues. Further, a western blot analysis revealed that a cytosolic protein of 110 kD, in both human colon and ileum, reacted with the monoclonal antibody to insulin-degrading enzyme. It is concluded that insulin-degrading enzyme is present in the cytosol of human colon and ileal mucosal cells.
Bai
J P
JP
College of Pharmacy, University of Minnesota, Minneapolis 55455, USA.
Hong
H J
HJ
Rothenberger
D A
DA
Wong
W D
WD
Buls
J G
JG
D000818:Animals; D015153:Blotting, Western; D018938:Caco-2 Cells; D003106:Colon / Q000187:drug effects / Q000201:enzymology*; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007082:Ileum / Q000187:drug effects / Q000201:enzymology*; D007328:Insulin / Q000378:metabolism*; D007329:Insulin Antagonists / Q000494:pharmacology*; D007339:Insulysin / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000187:drug effects / Q000201:enzymology; D051379:Mice
D004791:Enzyme Inhibitors; D007328:Insulin; D007329:Insulin Antagonists; D007339:Insulysin