RELATION OF ENZYME PROCESSING TO HEPATIC GLUCURONIDATION

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UDP-glucuronosyltransferases (UDPGTs) are a group of glycoprotein enzymes concentrated in the endoplasmic reticulum of liver cells, which are essential for the detoxification of internal metabolites and foreign substances, e.g. drugs, carcinogens and environmental toxins. UDPGTs exist as several isoforms; inherited defects of specific forms of UDPGT result in life-long jaundice in man and mutant (Gunn) rats. In preliminary studies we have shown that Gunn rats have approximately normal amounts of two functionally defective UDPGT isoforms. However, the biochemical basis of functional heterogeneity and inherited functional disorders of UDPGTs is not known. The purpose of the proposed project is to determine the possible role of glycosylation and other types of co-translational and post- translational changes in the generation of functional heterogeneity and inherited functional defects of UDPGTs. We plan to develop monoclonal and polyclonal antibodies that are specific for individual UDPGT isoforms and to use these to study the expression of UDPGT genes in inbred normal rats and congeneic Gunn rats. Precursors of individual UDPGT isoforms will be identified by cell-free translation of liver mRNA from normal rats; these precursors will be compared with those derived by translation of Gunn rat liver mRNA. Biosynthetic steps of synthesis of UDPGT precursors and processing of their protein and carbohydrate components will be studied by metabolic labeling of hepatocytes isolated from livers of Gunn rats and congeneic normal rats. Since UDPGTs are integral endoplasmic reticulum proteins, in addition to elucidation of the biochemical basis of inherited disorders of this essential enzyme system, these studies may help us understand the mechanism of assembly of liver endoplasmic reticulum proteins in general.