INFLUENCE OF GLUTATHIONE-S-TRANSFERASE-B (LIGANDIN) ON THE INTERMEMBRANE TRANSFER OF BILIRUBIN - IMPLICATIONS FOR THE INTRACELLULAR TRANSPORT OF NONSUBSTRATE LIGANDS IN HEPATOCYTES

To examine the hypothesis that glutathione S-transferases (GST) play a n important role in the hepatocellular transport of hydrophobic organi c anions, the kinetics of the spontaneous transfer of unconjugated bil irubin between membrane vesicles and rat liver glutathione S-transfera se B (ligandin) was studied, using stopped-flow fluorometry, Bilirubin transfer from glutathione S-transferase B to phosphatidylcholine vesi cles was best described by a single exponential function, with a rate constant of 8.0+/-0.7 s(-1) (+/-SD) at 25 degrees C. The variations in transfer rate with respect to acceptor phospholipid concentration pro vide strong evidence for aqueous diffusion of free bilirubin. This fin ding was verified using rhodamine-labeled microsomal membranes as acce pters. Bilirubin transfer from phospholipid vesicles to GST also exhib ited diffusional kinetics. Thermodynamic parameters for bilirubin diss ociation from GST were similar to those for human serum albumin. The r ate of bilirubin transfer from rat liver basolateral plasma membranes to acceptor vesicles in the presence of glutathione S-transferase B de clined asymptotically with increasing GST concentration. These data su ggest that glutathione S-transferase B does not function as an intrace llular bilirubin transporter, although expression of this protein may serve to regulate the delivery of bilirubin, and other nonsubstrate li gands, to sites of metabolism within the cell.