The ATP-depleting reagent "iodacetamide" induces the degradation of protein kinase C alpha (PKCα) in LLC-PK₁ pig kidney cells

The alkylating reagent iodoacetamide, a potent inhibitor of sulfhydryl proteases, was found to stimulate the selective degradation of protein kinase C alpha (PKCα) isoform. Treatment of LLC-PK₁ cells with iodoacetamide (5 mM) for 30-90 minutes at room temperature followed by western blotting on total cell homogenate, revealed the absence of an 80 KDa protein and the appearance of an 47 KDa band that was still recognized with the antibody. Serine protease inhibitor, metalloprotease inhibitors and leupeptin failed to prevent the degradation of PKCα. The degradation persisted at 4 °C and in the absence of Ca²⁺. Iodoacetamide had no direct effect on purified PKCα. In conclusion, the degradation of PKCα is a novel phenomenon. The degradation process could not be inhibited by known protease inhibitors or in the absence of Ca²⁺ or at 4 °C and appears to involve interactions with an unknown intermediates.