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. 2014 May 14;5(7):736–741. doi: 10.1021/ml5000405

Figure 2.

Figure 2

RXRα antagonist 23 binds RXRα at a site other than LBP. (A) Dose-dependent effect of 23 on inhibiting RXRα transactivation. HEK293T cells cotransfected with pG5-Luc and pBind-RXRα-LBD were treated with 9-cis-RA (10–-7 M) alone or together with the indicated concentration of 23 for 18 h. (B) SPR assay. Gradient concentrations of 23 were injected through flow cells immobilized with RXRα-LBD. The kinetic profiles are shown. The dissociation constant (Kd) of the 23/RXRα-LBD complex was calculated to be 4.881 × 10–7 M. (C) Compound 23 fails to compete with 9-cis-RA for binding to RXRα LBP. The bacterially expressed His-tagged RXRα-LBD was incubated with 7.5 nM [3H]-9-cis-RA in the presence or absence of the indicated concentrations of 9-cis-RA or 23. The RXRα LBD was captured by nickel-coated beads. Bound [3H]-9-cis-RA was quantitated by liquid scintillation counting. Data shown are mean ± SD.