Hemin inhibits ATP-dependent ubiquitin-dependent proteolysis: Role of hemin in regulating ubiquitin conjugate degradation

Abstract

Hemin has been shown to inhibit specifically the energy-dependent degradation of normal and abnormal proteins in reticulocytes. The present work demonstrates that the action of hemin involves the multi-enzyme ATP-dependent ubiquitin-dependent proteolytic system exclusively. At a concentration of ≃25 μM, hemin produces 50% inhibition of the degradation of 125I-labeled bovine serum albumin by this pathway. Hemin has no effect on the basal rate of proteolysis in the absence of either ATP or ubiquitin. At a concentration of hemin that gives complete inhibition of proteolysis, ATP-dependent formation of ubiquitin conjugates continues at about 50% of the control rate but the degradation of these ubiquitin conjugates is completely blocked. Inhibition of overall proteolysis and conjugate degradation are sensitive to hemin concentration to exactly the same extent. Hemin inhibition of conjugate breakdown results in the accumulation of higher molecular weight conjugates that are lost when hemin is removed by dilution. A model is proposed in which hemin acts as a negative allosteric effector in the initial step of a sequential degradative path by which intact ubiquitin conjugates are first cleaved to ubiquitin-associated fragments.