Fig. 5

NMR detected exchange between two long-lived T0070907-bound conformations. a Overlay of 2D [¹H,¹⁵N]-TROSY-HSQC NMR spectra of ¹⁵N-labeled PPARγ LBD bound to GW9662 or T0070907. b Binding of T0070907 but not GW9662 stabilizes intermediate exchange (µs-ms time scale) dynamics (residues labeled in a shown in green spheres) and causes peak doubling (tan and pink spheres; G399 is colored pink for emphasis). Data plotted on the T0070907-bound PPARγ crystal structure and important structural regions are highlighted as follows: AF-2 surface (black oval); an extended pyridyl-water hydrogen bond network (blue spheres, yellow dotted lines, blue oval), beyond the key pyridyl-water interaction (red sphere). c Snapshot overlays of 2D [¹H,¹⁵N]-TROSY-HSQC spectra of ¹⁵N-labeled PPARγ LBD bound to T0070907 or GW9662 shows co-linear shifting of peaks at the different temperatures. The spectral region displayed shows peaks conserved when PPARγ is bound to GW9662 or T0070907 (green arrows); a unique set of doubled peaks when bound to T0070907 (purple arrows); and absent peaks due to intermediate exchange on the NMR time scale when bound to GW9662 (dotted rectangles). d Snapshots of ZZ-exchange [¹H,¹⁵N]-HSQC NMR spectra (delay = 1 s) of T0070907-bound ¹⁵N-labeled PPARγ LBD focused on G399 at the indicated temperatures. Two G399 conformational states are denoted as A and B with the ZZ-exchange transfer crosspeaks as A–B and B–A. e ZZ-exchange NMR analysis build-up curve from for G399 at 310 K generated by plotting peak intensities of the state A and B peaks and exchange crosspeaks (A–B and B-A) as a function of delay time. f Schematic of the T0070907-bound PPARγ conformational ensemble defined by the NMR studies