The N-terminal regions of the estrogen receptor ␣ (ER␣-N) and ␤ (ER␤-N) were expressed and purified to homogeneity. Using NMR and circular dichroism spectroscopy, we conclude that both ER␣-N and ER␤-N are unstructured in solution. The TATA box-binding protein (TBP) has been shown previously to interact with ER␣-N in vitro and to potentiate ER-activated transcription. We used surface plasmon resonance and circular dichroism spectroscopy to confirm and further characterize The estrogen receptors (ERs)1 are ligand-inducible transcription factors that mediate the biological effects of estrogens. Two isoforms of human ER, encoded by two different genes, have been cloned and characterized, ER␣ and ER␤ (1-3). The ERs belong to a superfamily of nuclear receptors (NRs) that includes receptors for steroid hormones, thyroid hormones, and other hormones, as well as orphan receptors for which no ligand is known (4). All nuclear receptors share a similar modular structure; a variable N-terminal region, followed by a DNA binding domain (DBD), a hinge region, and a C-terminal ligand binding domain (LBD) (5). Transactivation regions have been localized to the LBD and in many cases also to the N-terminal region of the nuclear receptors (6, 7). The ligand-bound ERs bind as homodimers to specific DNA sequences termed estrogen response elements and regulate transcription through interaction with transcription modulators and recruitment of the general transcription machinery (8). ER␣ and ER␤ have also been shown to heterodimerize on estrogen response elements (9 -11).ER␣ contains two major transcription activation functions (AFs): one located in the N-terminal region (AF-1) and one in the C-terminal region of the LBD (AF-2) (12). A third activation function has been reported (AF-2a) residing in the boundary between the hinge and the LBD domains of ER␣ (13). The AF-1 function is hormone-independent, whereas the AF-2 function requires the presence of hormone (12). Full transcription activity of the ER␣ is thought to be achieved by synergism between the AFs. Further, the activities of the AFs are dependent on promoter and cellular context (14 -16). The ER␤ has a high homology to ER␣ in the DBD (96% amino acid identity) and in the LBD (58% amino acid identity) (2). In contrast, the Nterminal region of ER␤ is ϳ80 amino acids shorter than that of ER␣ and has also very poor sequence homology to that of ER␣. The N-terminal region in ER␤ is well conserved between different species such as rat, mouse, and human, which would imply functional importance.To date relatively little information has been available on the structure of the N-terminal regions of the NRs. The glucocorticoid receptor (GR) N-terminal transactivation region 1 (AF-1) and a shorter core fragment of 1, the 1core, have been shown by NMR and circular dichroism (CD) spectroscopy to be unstructured in aqueous solution (17). Furthermore, the isolated N-terminal transactivation region of the progesterone receptor A (PR A) has been shown to be sensitive to rapid degradation in limit...

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