Conjugation of ubiquitin to denatured hemoglobin is proportional to the rate of hemoglobin degradation in HeLa cells.

Publisher Website
Google Scholar
Abstract
Ubiquitin was radioiodinated and introduced into HeLa cells [human cervical carcinoma line] by the erythrocyte-mediated fusion procedure. Fractionation of injected HeLa cells and subsequent sodium dodecyl sulfate/polyacrylamide gel electrophoresis showed that HeLa nuclei contained 2 major labeled proteins, ubiquitin and the histone H2A-uniquitin conjugate, protein A24. HeLa cytosol contained ubiquitin and a series of ubiquitin-protein conjugates of diverse MW. When injected HeLa cells were treated with phenylhydrazine to denature the cotransferred Hb, a series of prominent ubiquitin-globin conjugates appeared. The identity of these conjugates was established by microinjection experiments in which both proteins were labeled. At low doses of phenylhydrazine, the intracellular concentration of globin-ubiquitin conjugates was proportional to the rate of Hb degradation. This result, together with the observation that ubiquitin conjugation to globin was markedly enhanced by phenylhydrazine-induced denaturation of Hb, provided support for the hypothesis that the covalent attachment of ubiquitin to proteins signals proteolysis.