Fragment screening using X-ray crystallography was carried out to discover starting points for hit optimization for PaFabF. For this purpose, the DSi-Poised library containing 875 compounds and the Leeds-3D library of the XChem facility at the Diamond synchrotron were screened by soaking PaFabF C164Q crystals. (Due to a limited number of available crystals, only 102 out of the 126 compounds in the Leeds/3D library were used for soaking). This resulted in 973 crystals which could be harvested and mounted for diffraction data collection, and 824 of the obtained datasets could be auto processed. Subsequent analysis was carried out using pan-dataset density analysis as implemented in PanDDA to increase the detection limit of weakly binding fragments. This required that all datasets were processed to the same space group and that their resolution was as high as possible (ideally below 3.0 Å). The ground model was processed to C 1 2 1 (with the unit cell a=137 Å, b=65 Å, c=84 Å, α==90°, β=93°) and thus only datasets that could be processed to the same space group and had sufficient resolution were considered for further analysis. Based on these criteria, 600 datasets were loaded for analysis in PanDDA out of which 582 were accepted and successfully processed. Of these 582 datasets, 219 were flagged to contain at least one difference electron density blob that needed manual inspection resulting in 382 electron density difference blobs for manual inspection altogether. Ligands were placed in 66 blobs (belonging to 57 different crystals), while the electron density difference of the remaining 316 potential binding events was not sufficient to justify unambiguous placement of a ligand. After a few rounds of refinement, 48 datasets were finally considered of quality good enough for deposition in the PDB (Table ).