Passos Ana L S, Pinto Patrícia I S, Power Deborah M, Canario Adelino V M
Centre of Marine Sciences, CCMAR-CIMAR Associate Laboratory, University of Algarve, Gambelas, Faro, Portugal.
Ecotoxicol Environ Saf. 2009 Jul;72(5):1529-37. doi: 10.1016/j.ecoenv.2009.02.004. Epub 2009 Mar 19.
A yeast (Saccharomyces cerevisiae)-based assay was developed and tested with steroids and chemicals (mostly pesticides). The induction of beta-galactosidase activity was strictly dependent on the presence of seabream (Sparus aurata) betaa estrogen receptor (sbERbetaa) and substances known to have estrogenic activity. 17beta-Estradiol (E(2)) and diethylstilbestrol (DES), both agonists, were most active and the antagonist tamoxifen (TAM) was 14-fold less active than E(2). Among the chemicals tested bisphenol-A was most active, followed by pentachlorophenol and naphthalene. Ligand-binding assays with recombinant sbERbetaa and sbERalpha revealed that sbERbetaa binds E(2) with 6.5-fold higher affinity than sbERalpha, confirming the selection of a high sensitive receptor for the yeast assay. DES, ICI 182,780, estrone and TAM had higher relative binding affinity to E2 in sbERalpha than sbERbetaa, although there was no difference in IC50 for these steroids between the two receptors. These results reveal the usefulness of using the yeast-based receptor assay for detecting chemical interaction with steroid receptors from contaminated samples.
开发了一种基于酵母（酿酒酵母）的检测方法，并用类固醇和化学物质（主要是农药）进行了测试。β-半乳糖苷酶活性的诱导严格依赖于鲷鱼（金头鲷）β雌激素受体（sbERβa）和已知具有雌激素活性的物质的存在。17β-雌二醇（E₂）和己烯雌酚（DES）这两种激动剂活性最强，拮抗剂他莫昔芬（TAM）的活性比E₂低14倍。在所测试的化学物质中，双酚A活性最强，其次是五氯苯酚和萘。用重组sbERβa和sbERα进行的配体结合试验表明，sbERβa与E₂的结合亲和力比sbERα高6.5倍，这证实了为酵母检测选择了一种高灵敏度受体。DES、ICI 182,780、雌酮和TAM与sbERα中的E₂相比，对E₂的相对结合亲和力更高，尽管这两种受体对这些类固醇的半数抑制浓度（IC₅₀）没有差异。这些结果揭示了使用基于酵母的受体检测方法来检测受污染样品中化学物质与类固醇受体相互作用的有用性。
