Detection and Characterization of a Novel Hepatic 8 S Binding Protein for Benzo[ a]Pyrene Distinct from the Ah Receptor

Abstract

Using fractionation procedures such as sucrose gradient sedimentation and gel permeation chromatography, a novel cytosolic binding protein for benzo[ a]pyrene has been detected in liver of nonmammalian and mammalian species including human. This protein, called 8 S protein, cosediments with the Ah receptor after centrifugation in sucrose density gradient but can be separated from the Ah receptor and 4 S protein by gel permeation chromatography. Owing to its binding characteristics, the 8 S protein is clearly distinct from the Ah receptor. The [ 3H]BP binding parameters have been determined by saturation experiments. According to Scatchard and Woolf plots the K d are 268 nM and 138 nM for DBA/2 mouse and guinea pig 8 S proteins, respectively. B max are 248 and 840 pmol/mg for DBA/2 mouse and guinea pig 8 S proteins, respectively. Apparent molecular mass of 8 S protein, according to Stokes radius (5 ± 0.2 nm) and sedimentation coefficient, was estimated ≍ 170,000 ± 6000. After in vitro incubation of 8 S proteins with [ 3H]BP the charcoal, as well as the 4 S protein, exerts potent stripping effects on the [ 3H]BP binding. In contrast, after administration of [ 3H]BP to DBA/2 mice the 8 S protein-[ 3H]BP complex formed in vivo is more resistant to the stripping effects of charcoal and 4 S protein. Competition studies demonstrate that polycyclic aromatic hydrocarbons (PAHs) (BP > 1-aminopyrene > pyrene > 7,12-dimethylbenz[ a]anthracene > 3-methylcholanthrene > benzo[ a]pyrene > benzo[ g, h, i,]perylene) are the best ligands of the 8 S protein. In contrast, the 2,3,7,8-tetrachlorodibenzo- p-dioxin is a poor ligand of this protein. Phenobarbital, steroid hormones, and omeprazole don′t bind the 8 S protein. The at present unknown function of the 8 S protein and its role in the toxicology of PAHs are currently under investigation.