Examples of bound TMAO molecules. Red, oxygen; blue, nitrogen; yellow or wheat, carbon; light blue, symmetry molecule. Numbers indicate contact distances in Å. Red dashed lines, contacts of 3.0 Å or less. Yellow dashed lines, contacts of >3.0 Å. The maps show OMIT DELFWT/PHDELFWT electron density from REFMAC contoured at +3σ/−3σ (cyan/red) (Murshudov et al., 2011). (a–c) Examples from trigonal trypsin at 1.75 Å resolution. The maps were calculated after ten cycles of restrained refinement of the final model with the TMAO molecules removed. (d) Example from orthorhombic trypsin at 1.70 Å resolution. The map was calculated after restrained refinement of the initial model with waters in the vicinity of the TMAO molecules removed but before building TMAO molecules into the model. This figure was prepared with PyMOL (Schrödinger LLC).