UBIQUITIN-MEDIATED DEGRADATION OF TYROSINE AMIMOTRANSFERASE (TAT) IN-VITRO AND IN-VIVO

Degradation of a protein via the ubiquitin proteolytic pathway involve s two successive steps. Covalent attachment of ubiquitin to the target protein and degradation of the tagged substrate by the 26S proteasome . Most native cellular proteins that are targeted by the ubiquitin sys tem are short-lived transcriptional activators and growth and cell cyc le regulators, as well as unstable membrane proteins. In the present s tudy we demonstrate the involvement of the system in the degradation o f tyrosine aminotransferase (TAT), a key enzyme in intermediary metabo lism. In vitro, we have shown that the native enzyme is conjugated and degraded in a system that requires ATP and ubiquitin. Degradation was monitored by following the decrease of catalytic activity as well as disappearance of the protein molecule. The enzyme could be protected f rom degradation by association with its specific cofactor, pyridoxal p hosphate (PLP). In vivo, we prepared cell extracts from livers of anim als in which TAT was induced by starvation and corticosteroid administ ration. The dramatic increase in the level of the enzyme was accompani ed by a concomitant increase in the level of specific TAT-ubiquitin ad ducts.