TY - JOUR
T1 - The RXRα C-terminus T462 is a NMR sensor for coactivator peptide binding
AU - Lu, Jianyun
AU - Chen, Minghe
AU - DeKoster, Gregory T.
AU - Cistola, David P.
AU - Li, Ellen
N1 - Funding Information:
We thank Mr. John Monsey and Dr. Alexander Koslov for assistance in ITC. This work was supported by grants from the National Institutes of Health (NIH) DK59501 (E. Li), Washington University Digestive Diseases Research Core Center (DK 52574), a pilot/feasibility award (J. Lu) from the Washington University Center for Human Nutrition (DK 56341). Spectra were collected at the Washington University Molecular Biophysics NMR laboratory and the High Resolution NMR Service Facility in Chemistry, which are funded in part by the NIH through NCRR shared Instrument Grants RR13874 and RR015715, respectively.
PY - 2008/2/22
Y1 - 2008/2/22
N2 - 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 receptor-interacting 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.
AB - 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 receptor-interacting 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.
KW - Activation
KW - Coactivator
KW - GRIP1
KW - Isothermal titration calorimetry
KW - NMR
KW - RXRα
UR - http://www.scopus.com/inward/record.url?scp=37549005189&partnerID=8YFLogxK
U2 - 10.1016/j.bbrc.2007.12.051
DO - 10.1016/j.bbrc.2007.12.051
M3 - Article
C2 - 18088598
AN - SCOPUS:37549005189
VL - 366
SP - 932
EP - 937
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
IS - 4
ER -