Topspin versions 2 1 3 2 and 3

NMR spectra were recorded at 298 K using a Bruker Avance III 700 MHz or 800 MHz spectrometers equipped with a triple resonance gradient standard probe or cryoprobe, respectively. Topspin versions 2.1, 3.2 and 3.5 (Bruker BioSpin) were used for data collection. Spectra processing used NMRPipe (35 (link)) followed by analysis with Sparky 3 (T.D. Goddard and D.G. Kneller, University of California) or NMRviewJ 8.0 (One Moon Scientific). Spectra for the assignment of backbone ¹H^N, ¹H^α, ¹³C’, ¹³C^α, ¹³C^β and ¹⁵N^H nuclei of full-length FKBP25 were collected on a 200 μM ¹³C,¹⁵N-labeled sample in 20 mM sodium phosphate (pH 6.5) with 150 mM NaCl, and 10% D₂O added for the lock. NMR spectra include 2D ¹⁵N-HSQC, 3D HNCO, 3D HNCACO, 3D HNCA. 3D HNCOCA, 3D CBCACONH, and 3D HNCACB. ¹H^N and ¹⁵N^H assignments for the full-length protein were used to assign ¹⁵N-HSQC spectra recorded for isolated BTHB and FKBP domains in the same buffer (20 mM sodium phosphate, pH 6.5, with 150 mM NaCl). For the BTHB domain, the assignments were confirmed by measuring additional 3D HNCO and 3D HNCACB spectra on a 310 μM sample of ¹³C,¹⁵N-labeled protein. Assignments of asparagine and glutamine sidechain amides were confirmed by using a 3D ¹⁵N-HSQC-NOESY spectrum (150 ms mixing time).