2.1. Protein purification, crystallization and structure determination  

Purification and crystallization of TgPRS was performed in accordance with previously published methods (Jain et al., 2014 ▸, 2015 ▸; Jain, Yogavel et al., 2017 ▸). Diffraction-quality crystals of apo TgPRS were obtained in Morpheus (Molecular Dimensions) condition D3, while crystals of the TgPRS–FF–AMPPNP complex grew in Morpheus II (Molecular Dimensions) condition E9. Diffraction data were collected on the PROXIMA 1 and 2A beamlines at Synchrotron SOLEIL, France. A total of 3600 frames were collected in 0.1° oscillation steps with 0.1 s exposure per frame. The data were processed and scaled using DIALS (Winter et al., 2018 ▸) and the relevant statistics are summarized in Table 1 ▸. The structures were solved by molecular replacement, with apo TgPRS (PDB entry 5xif; Jain, Yogavel et al., 2017 ▸) as a search model for the apo structure. The structure of TgPRS–HF–AMPPNP (PDB entry 5xiq; Jain, Yogavel et al., 2017 ▸) was used as a search model for TgPRS–FF–AMPPNP. All structures were solved with Phenix (Adams et al., 2010 ▸). X-ray refinement restraint parameters were generated for the ligand febrifugine using the grade web server (http://grade.globalphasing.org). These were subsequently refined using phenix.refine. NCS was not used during refinement, but TLS was. The atomic structures were subjected to refinement cycles using simulated annealing (Cartesian) for three cycles. This was used to remove model bias. After each step, the models were manually adjusted so there was coherence in 2F _o − F _c and F _o − F _c electron-density maps. Coot (Emsley et al., 2010 ▸) was used for all rebuilding. The presence of the ligands was confirmed using OMIT maps (Fig. 1 ▸). Model quality was assessed using the MolProbity server (Chen et al., 2010 ▸). Atomic cooordinates and structure factors have been deposited in the PDB under accession codes 6aa0 and 6a88.

Electron-density fit of FF and AMPPNP. A composite OMIT difference Fourier map (F _o – F _c) generated at 2.6 Å and contoured at 1.7σ showing bound FF (coral), AMPPNP (dark green) and the single Mg²⁺ ion (cyan).

Values in parentheses are for the highest resolution shell.