Hello all, I’m an undergraduate working on structure calculation for a protein – ligand complex for months but haven’t got any satisfying results so far. The following is my protocol and criteria for X-ray diffraction data processing.

(1) Process the diffraction data with HKL-2000 and obtain output file. (*.sca)

(2) Run Xtriage in PHENIX to search for any error. Only if the Xtriage result shows no red light, go on to the Phaser-MR (simple one-component interface.)

(3) Top LLG should be positive and as high as possible, while Top TFZ should be more than 8 or 6. When Phaser-MR result says "No MR solution found", change search options and run again. (Fig. 1)

(4) Refine the data with Phenix.refine. Select Rigid body, Use NCS, and Simulated annealing (Cartesian) options in Refinement settings on the first refinement. Select Update water additively if it’s more than second refinement. (Fig. 2) The resultant R-work and R-free should be around 0.2.

(5) Open *.mtz and *.pdb in WinCoot. Start Stepped refine and Refine/improve Ramachandran plot.

(6) Save coordinates.

Recently, I found out that two MTZ file - *.mtz and *_data.mtz - are created after refinement in step (4). The problem is, I’ve been using *.mtz in refinement rather than *_data.mtz. Could this be a problem for something? And if possible, can you advise me for the protocol I’m following?

Best regards.