Insulin-degrading enzyme in a human colon adenocarcinoma cell line (Caco-2).
The activity of insulin-degrading enzyme (IDE), a thiol metalloprotease degrading insulin in many insulin target cells, was determined in human colon adenocarcinoma (Caco-2) cells. Insulin-degrading activity was localized in the cytosol of Caco-2 cells, accounting for 88% of total activity. Western blots and immunoprecipitation showed that IDE was present in the cytosol of Caco-2 cells and contributed to more than 93% cytosolic insulin-degrading activity. Cytosolic insulin degradation was strongly inhibited by IDE inhibitors, including N-ethylmaleimide, 1,10-phenanthroline, p-chloromericuribenzoate, and EDTA, but was not significantly or not as extensively inhibited by strong inhibitors of proteasome, i.e., chymostatin, soybean trypsin inhibitor, leupeptin, and Dip-F. These results suggest that IDE is present in Caco-2 cells, that Caco-2 IDE has properties similar to those of its counterparts in insulin-target tissues, and that it significantly contributes to intracellular insulin degradation.
Bai
J P
JP
Department of Pharmaceutics, College of Pharmacy, University of Minnesota, Minneapolis 55455, USA.
Hsu
M J
MJ
Shier
W T
WT
D000230:Adenocarcinoma / Q000201:enzymology*; D003110:Colonic Neoplasms / Q000201:enzymology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007339:Insulysin / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D007339:Insulysin