To the Phenixbb community: I've been experimenting with using TLS to refine my 2.4 angstrom resolution structure. There are 2 protomers in the ASU, and each has a metal cluster and a small-molecule ligand. The model has a relatively high average B-factor for protein (~65) atoms and I also suspect that both the cluster and the ligand have partial occupancy. At this resolution however, I've modeled both groups with 100% occupancy and the average B-factors for the atoms of these groups are expectedly higher, ~100 after individual_sites and individual_adp refinement (i.e. no TLS). My problem arises when I try to use TLS. I determined the boundaries for the TLS groups using the TLSMD server, which identified 3 groups per protomer (roughly coincide with domain boundaries). After including TLS refinement, the R and R-free both improve by 1-2%, but the B-factors for the clusters and ligands end up very high. Metal atoms in the cluster have B-factors >350 and the ligand atoms have B-factors >120. The average B-factor for protein atoms is slightly higher too (~70). Also, the gap between R and Rfree is larger than I'd expect (19% R and 26% Rfree), although this isn't unheard of and more importantly, the values have fully converged yet. So, should I be treating the ligands differently if I intend to use TLS? I tried including them with one of the TLS groups, but that didn't help. Any other ideas are welcome. Best Regards, -Andy Torelli