Limits...
Direct evidence revealing structural elements essential for the high binding ability of bisphenol A to human estrogen-related receptor-gamma.

Okada H, Tokunaga T, Liu X, Takayanagi S, Matsushima A, Shimohigashi Y - Environ. Health Perspect. (2008)

Bottom Line: When we examined BPA derivatives to evaluate the structural essentials required for the binding of BPA to ERR-gamma , we found that only one of the two phenol-hydroxyl groups was essential for the full binding.The maximal activity was attained when one of the methyl groups was removed.These results indicate that the phenol derivatives are potent candidates for the endocrine disruptor that binds to ERR-gamma.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Structure-Function Biochemistry, Department of Chemistry, The Research-Education Centre of Risk Science, Faculty and Graduate School of Sciences, Kyushu University, Fukuoka, Japan.

ABSTRACT

Background: Various lines of evidence have shown that bisphenol A [BPA; HO-C6H4-C(CH3)2-C6H4-OH] acts as an endocrine disruptor when present in very low doses. We have recently demonstrated that BPA binds strongly to human estrogen-related receptor-gamma (ERR-gamma ) in a binding assay using [3H]4-hydroxytamoxifen ([3H]4-OHT). We also demonstrated that BPA inhibits the deactivation activity of 4-OHT.

Objectives: In the present study, we intended to obtain direct evidence that BPA interacts with ERR-gamma as a strong binder, and also to clarify the structural requirements of BPA for its binding to ERR-gamma.

Methods: We examined [3H]BPA in the saturation binding assay using the ligand binding domain of ERR-gamma and analyzed the result using Scatchard plot analysis. A number of BPA derivatives were tested in the competitive binding assay using [3H]BPA as a tracer and in the luciferase reporter gene assay.

Results: [3H]BPA showed a KD of 5.50 nM at a Bmax of 14.4 nmol/mg. When we examined BPA derivatives to evaluate the structural essentials required for the binding of BPA to ERR-gamma , we found that only one of the two phenol-hydroxyl groups was essential for the full binding. The maximal activity was attained when one of the methyl groups was removed. All of the potent BPA derivatives retained a high constitutive basal activity of ERR-gamma in the luciferase reporter gene assay and exhibited a distinct inhibitory activity against 4-OHT.

Conclusion: These results indicate that the phenol derivatives are potent candidates for the endocrine disruptor that binds to ERR-gamma.

Show MeSH

Related in: MedlinePlus

Chemical structure of BPA and its derivatives and their dose–response curves in the radioligand receptor binding assay for ERR-γ . (A) Chemical structures of BPA (two methyl groups) and its derivatives: bisphenol B (a methyl group and an ethyl group) and bisphenol AP (a methyl group and a phenyl group). (B) Binding activities of BPA, bisphenol B, and bisphenol AP examined by the competitive binding assay using [3H]BPA and GST–ERR-γ –LBD. (C) Chemical structures of bisphenol E (one methyl group) and its derivatives, bisphenol F and bisphenol AF [two trifluoromethyl groups (CF3)]. (D) Binding activities of BPA, bisphenol E, bisphenol F, and bisphenol AP examined by the competitive binding assay. (B) and (D) each show representative curves with the IC50 values closest to the mean IC50 from at least five independent assays for each compound. B/B0 is the relative inhibitory activity estimated from the calculation of the percentage of displacement by the chemical tested (B) against the specific binding (B0 = 100%) of [3H]BPA.
© Copyright Policy - public-domain
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2199305&req=5

f2-ehp0116-000032: Chemical structure of BPA and its derivatives and their dose–response curves in the radioligand receptor binding assay for ERR-γ . (A) Chemical structures of BPA (two methyl groups) and its derivatives: bisphenol B (a methyl group and an ethyl group) and bisphenol AP (a methyl group and a phenyl group). (B) Binding activities of BPA, bisphenol B, and bisphenol AP examined by the competitive binding assay using [3H]BPA and GST–ERR-γ –LBD. (C) Chemical structures of bisphenol E (one methyl group) and its derivatives, bisphenol F and bisphenol AF [two trifluoromethyl groups (CF3)]. (D) Binding activities of BPA, bisphenol E, bisphenol F, and bisphenol AP examined by the competitive binding assay. (B) and (D) each show representative curves with the IC50 values closest to the mean IC50 from at least five independent assays for each compound. B/B0 is the relative inhibitory activity estimated from the calculation of the percentage of displacement by the chemical tested (B) against the specific binding (B0 = 100%) of [3H]BPA.

Mentions: We evaluated the role of the two methyl (CH3) groups on the sp3-C atom of BPA in binding to ERR-γ by a series of analogs of BPA, HO-C6H4-C(CH3)2-C6H4-OH. First, we examined the effect of incorporation of the methyl group on the binding affinity of BPA. When CH3 was incorporated into the parent methyl group to produce HO-C6H4-C(CH3)(CH2CH3)-C6H4-OH (Figure 2A), we found the resulting bisphenol B to be approximately half as potent (IC50 = 26.3 nM) as BPA (Table 1). This result clearly indicates that a bulky group on the central sp3-C atom is obviously disadvantageous in terms of the binding of BPA to ERR-γ ’s binding pocket.


Direct evidence revealing structural elements essential for the high binding ability of bisphenol A to human estrogen-related receptor-gamma.

Okada H, Tokunaga T, Liu X, Takayanagi S, Matsushima A, Shimohigashi Y - Environ. Health Perspect. (2008)

Chemical structure of BPA and its derivatives and their dose–response curves in the radioligand receptor binding assay for ERR-γ . (A) Chemical structures of BPA (two methyl groups) and its derivatives: bisphenol B (a methyl group and an ethyl group) and bisphenol AP (a methyl group and a phenyl group). (B) Binding activities of BPA, bisphenol B, and bisphenol AP examined by the competitive binding assay using [3H]BPA and GST–ERR-γ –LBD. (C) Chemical structures of bisphenol E (one methyl group) and its derivatives, bisphenol F and bisphenol AF [two trifluoromethyl groups (CF3)]. (D) Binding activities of BPA, bisphenol E, bisphenol F, and bisphenol AP examined by the competitive binding assay. (B) and (D) each show representative curves with the IC50 values closest to the mean IC50 from at least five independent assays for each compound. B/B0 is the relative inhibitory activity estimated from the calculation of the percentage of displacement by the chemical tested (B) against the specific binding (B0 = 100%) of [3H]BPA.
© Copyright Policy - public-domain
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2199305&req=5

f2-ehp0116-000032: Chemical structure of BPA and its derivatives and their dose–response curves in the radioligand receptor binding assay for ERR-γ . (A) Chemical structures of BPA (two methyl groups) and its derivatives: bisphenol B (a methyl group and an ethyl group) and bisphenol AP (a methyl group and a phenyl group). (B) Binding activities of BPA, bisphenol B, and bisphenol AP examined by the competitive binding assay using [3H]BPA and GST–ERR-γ –LBD. (C) Chemical structures of bisphenol E (one methyl group) and its derivatives, bisphenol F and bisphenol AF [two trifluoromethyl groups (CF3)]. (D) Binding activities of BPA, bisphenol E, bisphenol F, and bisphenol AP examined by the competitive binding assay. (B) and (D) each show representative curves with the IC50 values closest to the mean IC50 from at least five independent assays for each compound. B/B0 is the relative inhibitory activity estimated from the calculation of the percentage of displacement by the chemical tested (B) against the specific binding (B0 = 100%) of [3H]BPA.
Mentions: We evaluated the role of the two methyl (CH3) groups on the sp3-C atom of BPA in binding to ERR-γ by a series of analogs of BPA, HO-C6H4-C(CH3)2-C6H4-OH. First, we examined the effect of incorporation of the methyl group on the binding affinity of BPA. When CH3 was incorporated into the parent methyl group to produce HO-C6H4-C(CH3)(CH2CH3)-C6H4-OH (Figure 2A), we found the resulting bisphenol B to be approximately half as potent (IC50 = 26.3 nM) as BPA (Table 1). This result clearly indicates that a bulky group on the central sp3-C atom is obviously disadvantageous in terms of the binding of BPA to ERR-γ ’s binding pocket.

Bottom Line: When we examined BPA derivatives to evaluate the structural essentials required for the binding of BPA to ERR-gamma , we found that only one of the two phenol-hydroxyl groups was essential for the full binding.The maximal activity was attained when one of the methyl groups was removed.These results indicate that the phenol derivatives are potent candidates for the endocrine disruptor that binds to ERR-gamma.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Structure-Function Biochemistry, Department of Chemistry, The Research-Education Centre of Risk Science, Faculty and Graduate School of Sciences, Kyushu University, Fukuoka, Japan.

ABSTRACT

Background: Various lines of evidence have shown that bisphenol A [BPA; HO-C6H4-C(CH3)2-C6H4-OH] acts as an endocrine disruptor when present in very low doses. We have recently demonstrated that BPA binds strongly to human estrogen-related receptor-gamma (ERR-gamma ) in a binding assay using [3H]4-hydroxytamoxifen ([3H]4-OHT). We also demonstrated that BPA inhibits the deactivation activity of 4-OHT.

Objectives: In the present study, we intended to obtain direct evidence that BPA interacts with ERR-gamma as a strong binder, and also to clarify the structural requirements of BPA for its binding to ERR-gamma.

Methods: We examined [3H]BPA in the saturation binding assay using the ligand binding domain of ERR-gamma and analyzed the result using Scatchard plot analysis. A number of BPA derivatives were tested in the competitive binding assay using [3H]BPA as a tracer and in the luciferase reporter gene assay.

Results: [3H]BPA showed a KD of 5.50 nM at a Bmax of 14.4 nmol/mg. When we examined BPA derivatives to evaluate the structural essentials required for the binding of BPA to ERR-gamma , we found that only one of the two phenol-hydroxyl groups was essential for the full binding. The maximal activity was attained when one of the methyl groups was removed. All of the potent BPA derivatives retained a high constitutive basal activity of ERR-gamma in the luciferase reporter gene assay and exhibited a distinct inhibitory activity against 4-OHT.

Conclusion: These results indicate that the phenol derivatives are potent candidates for the endocrine disruptor that binds to ERR-gamma.

Show MeSH
Related in: MedlinePlus