Structure-guided drug discovery emerged in the 1970s and 1980s, stimulated by the 3D-structures of protein targets that became available, mainly through X-ray crystal structure analysis, assisted by development of synchrotron radiation sources. Structures of known drugs or inhibitors were used to guide the development of leads. The growth of high-throughput screening during the late 1980s and the early 1990s in the pharmaceutical industry of chemical libraries of hundreds of thousands of compounds of molecular weight of ~500 Da was impressive, but still explored only a tiny fraction of the chemical space of the predicted 1040 drug-like compounds. The use of fragments with molecular weights less than 300 Da in drug discovery, not only decreased the chemical space needing exploration but also increased promiscuity in binding targets. Here we discuss advances in X-ray fragment screening and the challenge of identifying sites where fragments not only bind but can be chemically elaborated while retaining their positions and binding modes. We first describe the analysis of fragment binding using conventional X-ray difference Fourier techniques, using M. abscessus SAICAR synthetase (PurC) as an example. We observe that all fragments occupy positions predicted by computational hotspot mapping. We compare this with fragment screening at Diamond Synchrotron Light Source XChem facility using PanDDA software, which identifies many more fragment hits, only some of which bind to the predicted hotspots. Many low occupancy sites identified may not support elaboration to give adequate ligand affinity, although they will likely be useful in drug discovery as “warm spots” for guiding elaboration of fragments bound at hotspots. We discuss the implications of these observations for fragment screening at the synchrotron sources. / The Botnar Foundation (grant number: 6063), the Cystic Fibrosis Trust (Strategic Research Centre Awards 002, 010 & 201) and the Bill and Melinda Gates Foundation, Shorten-TB Award.