ISOZYMES OF UBIQUITIN PEPTIDYL TRANSFERASE (PROTEIN DEGRADATION)
LEE, PAULINE LING
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ATP dependent protein degradation in reticulocyte lysate requires ubiquitin, a highly conserved heat stable protein. In the presence of ATP, ubiquitin is found to form alkaline-stable covalent conjugates with protein substrates. Ubiquitin conjugates are thought to play a critical role in protein degradation. It was first proposed that the covalent attachment of ubiquitin targets protein substrates for proteolytic degradation. However, more recent evidence suggests an alternative model where ubiquitin derepresses a protease by inactivating its endogenous inhibitor.;Ubiquitin conjugation requires activation of ubiquitin to a high energy intermediate by ubiquitin activating enzyme. It has been postulated that activated ubiquitin is then transferred to lysine residues of protein substrates through the action of ubiquitin conjugating enzyme or ubiquitin peptidyl transferase.;We isolated ubiquitin peptidyl transferases from human erythrocytes. Four fractions of ubiquitin peptidyl transferase activity were resolved. Each transferase fraction catalyzed the binding of a single ubiquitin molecule onto each substrate molecule. The transferase fractions differed in their specificities toward added substrates. A substrate for one transferase fraction was not necessarily a substrate for another transferase fraction. Two of the transferase fractions exhibited activity toward only a narrow range of substrates. In addition, a pair of endogenous proteins were found which are highly efficient substrates for ubiquitin conjugation.;The substrate specificities of the red cell ubiquitin peptidyl transferases that we studied suggest that not all proteins are substrates for ubiquitin conjugation but rather that specific intracellular protein substrates for ubiquitin conjugation exist. Thus, it appears that ubiquitin conjugation is not involved in targeting protein substrates for proteolysis but is involved in regulating the activities of specific intracellular proteins.