Polychlorinated Biphenyls (PCBs) Exhibit Differential Binding To Recombinant Human And Rainbow Trout Estrogen Receptors

Polychlorinated biphenyls (PCBs) exhibit differential binding to recombinant human and rainbow trout estrogen receptors. JB Matthews and TR Zacharewski.

Although the function and activities of estrogen and the ER are conserved between species, the amino acid sequence of the regions involved in ligand binding (D, E and F domains) are poorly conserved. This suggests that species may exhibit different responses and sensitivities to non-traditional ligands and that one species may not be an appropriate surrogate to identify and predict responses in other species. In order to investigate potential species-specific sensitivities to non-traditional ER ligands, a comparative study was undertaken in which we examined the ability of 20 structurally distinct PCBs (IUPAC #s: 41, 51, 58, 60, 68, 78, 91, 99, 104, 112, 115, 126, 143, 153, 169, 173, 184, 188, 190, 193) at concentrations ranging from 10-9 to 10-5 M to compete with [³H]-17b -estradiol (E2) for binding to bacterially expressed GST-human ERDEF (GST-hERdef; amino acids 264-595) and GST-rainbow trout ER DEF (GST-rtERdef; amino acids 224-575) fusion proteins. Both fusion proteins were purified to greater than 90% and yields provided sufficient GST-hERdef and GSTrtERdef fusion protein for approximately 300 and 100 binding assays, respectively, using a 96-well high throughput assay. The GST-hERdef and GST-rtERdef both exhibited high affinity for E2 with dissociation constants (K_d) of 1.28 ± 0.10 nM and 2.14 ± 0.30 nM, respectively. Of the 20 PCBs tested, PCB184 (2,2',3,4,4',6,6'-heptaCB; IC₅₀ = 0.51 µM), PCB104 (2,2',4,6,6'-pentaCB; IC50 = 0.83 µM) and PCB188 (2',3,4',5,6,6'-heptaCB; IC₅₀ = 1.3 µM) effectively competed with [3H]E2 for binding to the GSThERdef fusion protein. Conversely, six PCBs competed with [3H]E2 for binding to the GSTrtERdef fusion protein: PCB184 (IC₅₀ = 0.92 µM), PCB104 (IC₅₀ = 1.8 µM), PCB188 (IC₅₀ = 3.8 µM), PCB51 (2,2',4,6'-tetraCB; IC₅₀ = 5.5 µM), PCB91 (2,2',4,4'6-pentaCB IC₅₀ = 6.7 µM), PCB41 (2,2',3,4-tetraCB IC₅₀ = 9.5 µM). This data demonstrates that ERs from different species exhibit differential ligand preferences and affinities, and that the use of a single surrogate model may be unsuitable to predict estrogenic responses in all species.