The N-terminal Regions of Estrogen Receptor α and β Are Unstructured in Vitro and Show Different TBP Binding Properties

• Published in: The Journal of biological chemistry 2001-12, Vol.276 (49), p.45939-45944
• Main Authors: Wärnmark, Anette, Wikström, Anja, Wright, Anthony P.H., Gustafsson, Jan-Åke, Härd, Torleif
• Format: Article
• Language: English
• Subjects: Base Sequence; Cell Line, Transformed; Circular Dichroism; DNA Primers; DNA-Binding Proteins - metabolism; Electrophoresis, Polyacrylamide Gel; Estrogen Receptor alpha; Estrogen Receptor beta; In Vitro Techniques; Medicin och hälsovetenskap; Nuclear Magnetic Resonance, Biomolecular; Protein Binding; Receptors, Estrogen - chemistry; Receptors, Estrogen - metabolism; Surface Plasmon Resonance; TATA-Box Binding Protein; Transcription Factors - metabolism
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M107875200

The N-terminal regions of the estrogen receptor α (ERα-N) and β (ERβ-N) were expressed and purified to homogeneity. Using NMR and circular dichroism spectroscopy, we conclude that both ERα-N and ERβ-N are unstructured in solution. The TATA box-binding protein (TBP) has been shown previously to interact with ERα-N in vitro and to potentiate ER-activated transcription. We used surface plasmon resonance and circular dichroism spectroscopy to confirm and further characterize the ER-N-TBP interaction. Our results show that the intrinsically unstructured ERα-N interacts with TBP, and suggest that structural changes are induced in ERα-N upon TBP interaction. Conformational changes upon target factor interaction have not previously been demonstrated for any N-terminal region of nuclear receptors. In addition, no binding of ERβ-N to TBP was detected. This difference in TBP binding could imply differential recruitment of target proteins by ERα-N and ERβ-N. The affinity of the ERα-N-TBP interaction was determined to be in the micromolar range (KD = 10−6 to 10−5m). Our results support models of TBP as a target protein for the N-terminal activation domain of ERα. Further, our results suggest that target proteins can induce and/or stabilize ordered structure in N-terminal regions of nuclear receptors upon interaction.