The long-term objectives of this research is to provide better understanding of the causes and treatment of unconjugated hyperbilirubinemia in man. During neonatal life and in certain individuals with the Crigler-Najjar Syndrome, Types 1 and 2, the concentrations of bilirubin in the circulation can reach levels that are associated with bilirubin encephalopathy. The basis for this disturbance is believed due to a delayed appearance or genetic absence of the glucuronyl transferase responsible for conjugation of bilirubin in liver microsomes. The Gunn rat is an animal model for this deficiency. The focus of the grant is the isolation of the normal and abnormal transferase enzyme from Gunn rat liver. The enzyme will be solubilized from liver microsomes, subjected to ammonium sulfate fractionation and purified by anion exchange, gel filtration and affinity chromotography. Amino acid and phospholipid composition studies are to be done on the normal and defective transferase. Normal and synthetic substrates will be used to better characterize the active site on the enzyme protein. The synthetic substrates are to be purified by high performance liquid chromatography as will the conjugation products. In vitro attempts will be made to correct the abnormal transferase from the jaundiced rat by both protein phosphorylations and dephosphorylations. Eventually the appearance and maturation of this transferase enzyme will be studied in fetal, neonatal and suckling animals.
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