This project is directed toward understanding the role of the Ubiquitin System in cellular metabolism. Reagents for immunochemical studies of Ubiquitin and Ubiquitin Activating Enzyme were developed and used for quantitation of these two proteins in Normal and Alzheimer's Diseased brains. Slightly lower levels of Ubiquitin Activating Enzyme were detected in Alzheimer's Diseased tissue. Immunocytochemical studies were carried out with rat brain, heart, liver and kidney and with HeLa cells. Results suggest significant compartmentalization of Ubiquitin Activating Enzyme in the cell nucleus. Two isoforms of Ubiquitin Activating Enzyme have been detected in several cell types. Characterization of these isoforms is in progress. Copurification of Ubiquitin Activating Enzyme and Ubiquitin Carrier Protein from rabbit reticulocytes has been observed, and may be an indicator of a specific protein-protein complex. Such an interaction may be the basis of a regulatory mechanism. Confirming evidence for complex formation is being sought.
