The unusual binding properties of the third distinct teleost estrogen receptor subtype ERbetaa are accompanied by highly conserved amino acid changes in the ligand binding domain.

Three forms of estrogen receptor: ERalpha, ERbeta (ERbetab), and a second ERbeta, ERbetaa (formerly ERgamma) are present in teleost fish. All ERbetaas share amino acid changes in the ligand binding domain that may influence ligand specificity and receptor function. We compared binding specificities of the three ERs of the teleost fish, Atlantic croaker Micropogonias undulatus. Bacterially expressed Atlantic croaker (ac) ERalpha, -betab, and -betaa fusion proteins showed specific, high affinity binding to 17beta-[(3)H]estradiol, with K(d) values of 0.61 +/- 0.013, 0.40 +/- 0.006, and 0.38 +/- 0.059 nm, respectively. Rank orders of binding were: diethylstilbestrol >> ICI182780 > 4-hydroxytamoxifen > ICI164384 > estradiol >/= zearalenone > moxestrol > tamoxifen > estrone >/= 17alpha-estradiol > estriol > 2-hydroxyestrone = genistein >> RU486 for acERalpha; ICI182780 > diethylstilbestrol > 4-hydroxytamoxifen > estradiol > ICI164384 > genistein > moxestrol > tamoxifen > zearalenone = estrone > estriol = 17alpha-estradiol > 2-hydroxyestrone >> RU486 for acERbetab; and estradiol >/= diethylstilbestrol > 4-hydroxytamoxifen > ICI182780 > ICI 164384 > estriol >/= genistein > moxestrol > zearalenone > estrone > 17alpha-estradiol > RU486 >/= tamoxifen > 2-hydroxyestrone for acERbetaa. acERbetaa showed higher relative binding affinities for estradiol, estriol, and RU486 and lower relative binding affinities for synthetic estrogens and antiestrogens than previously characterized ERs. Mutation of the conserved teleost substitutions (acERbetaaPhe(396)) to the ERalpha or ERbetab counterpart shifted diethylstilbestrol and tamoxifen affinities toward those of wild-type acERalpha and acERbetab, supporting the hypothesis that the positions with conserved residue changes in teleost ERs are important to ER structure and function.