The RXR-alpha C-terminus T462 is a NMR sensor for coactivator peptide binding
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Date
2008-02-22
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Authors
Lu, Jianyun
Chen, Minghe
DeKoster, Gregory T.
Cistola, David
Li, Ellen
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East Carolina University
Abstract
The C-terminal activation function-2 (AF-2) helix plays a crucial role in retinoid X receptor alpha (RXRα)-mediated gene expression. Here, we report a nuclear magnetic resonance (NMR) study of the RXRα ligand-binding domain complexed with 9-cis-retinoic acid and a glucocorticoid receptorinteracting protein 1 peptide. The AF-2 helix and most of the C-terminal residues were undetectable due to a severe line-broadening effect. Due to its outstanding signal-to-noise ratio, the C-terminus
residue, threonine 462 (T462) exhibited two distinct crosspeaks during peptide titration, suggesting that peptide binding was in a slow exchange regime on the chemical shift timescale. Consistently,
the Kd derived from T462 intensity decay agreed with that derived from isothermal titration calorimetry. Furthermore, the exchange contribution to the 15N transverse relaxation rate was measurable in either T462 or the bound peptide. These results suggest that T462 is a sensor for
coactivator binding and is a potential probe for AF-2 helix mobility. Originally published Biochemical and Biophysical Research Communications, Vol. 366, No. 4, Feb 2008
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Biochemical and Biophysical Research Communications; 366:4 p. 932-937