Setting up the JSON project file

Enter the main directory.

Note

You can find a ready-to-use JSON template file named

elongator_refine_template.json

in the tutorial directory and immediately proceed to the next step of Setting up the parameter file.

Generating the file from scratch

1. Generate the template for the refinement.

   gen_refinement_template.py --out_json elongator_refine_template.json --params params.py elongator.json
   

2. In the following steps, modify the resulting elongator_refine_template.json template file by adding new restraints and inactivating or removing restraints not needed, adjusting weights.

3. Add the traditional cross-correlation-based EM fit restraint to the “data” block in the JSON file:

   {
       "series": [
   
       ],
       "symmetry": {
           "sym_tr3ds": [
   
           ]
       },
       "data": [
           {
               "active": true,
               "type": "em_map",
               "name": "FitRestraint",
               "em_restraint_type": "FitRestraint",
               "filename": "EM_data/emd_4151_binned.mrc",
               "threshold": 0.01,
               "voxel_size": 6.6,
               "resolution": 25,
               "weight": 1000,
               "first_copy_only": true,
               "repr_resolution": 10,
               "optimized_components": [
                   {"name": "Elp1", "subunit": "Elp1"},
                   {"name": "Elp2", "subunit": "Elp2"},
                   {"name": "Elp3", "subunit": "Elp3"}
   
               ]
           }
       ],
       "subunits": [
   
       ],
       "xlinkanalyzerVersion": "0.1"
   }
   

4. Redefine rigid bodies. In this case, we noticed that some crosslinks are in long loops and tails. We will then let them move flexibly in the refinement.

   To to do this, we add rigid_bodies block to the JSON file, in which we define rigid bodies with resi_ranges, excluding the selected loops:

   {
       "series": [
   
       ],
       "rigid_bodies": [
           {
               "foreach_serie": true,
               "foreach_copy": true,
               "components": [
                   { "name": "Elp1_1", "subunit": "Elp1", "domain": "propeller1"}
               ]
           },
           {
               "foreach_serie": true,
               "foreach_copy": true,
               "components": [
                   { "name": "Elp1_2", "subunit": "Elp1", "domain": "propeller2"}
               ]
           },
           {
               "foreach_serie": true,
               "components": [
                   { "name": "Elp1_3", "subunit": "Elp1", "domain": "CTD", "copies": [0, 1]}
               ],
               "freeze": true
           },
           {
               "foreach_serie": true,
               "foreach_copy": true,
               "components": [
                   { "name": "Elp2", "subunit": "Elp2",
                       "resi_ranges": [
                           [1, 239],
                           [257, 788]
                       ]
                   }
               ]
           },
           {
               "foreach_serie": true,
               "foreach_copy": true,
               "components": [
                   { "name": "Elp3", "subunit": "Elp3",
                       "resi_ranges": [
                           [95, 468],
                           [477, 557]
                       ]}
               ]
           }
       ],
       "symmetry": {
           "sym_tr3ds": [
   
           ]
       },
       "data": [
   
       ],
       "subunits": [
   
       ],
       "xlinkanalyzerVersion": "0.1"
   }
   

   Note

   Whenever you define custom rigid bodies here, set add_rbs_from_pdbs = False in the parameter file.

5. Add symmetry restraint for the C-terminal domain of Elp1 (CTD).

   Why? The input structure for the CTD domain is already a symmetrical dimer and the entire dimer is defined as a rigid body. This domain cannot be constrained by the symmetry “constraint” because the constraints do not work “within rigid bodies”.

   To define the symmetry restraint, add the following block:

   {
       "series": [
   
       ],
       "rigid_bodies": [
   
       ],
       "symmetry": {
           "sym_tr3ds": [
   
           ]
           "apply_symmetry": [
               {
                   "sym": "2fold",
                   "restraint_type": "symmetry_restraint",
                   "selectors": [
                       {"subunit": "Elp1", "serie": "2fold", "copies": [0], "domain": "CTD"},
                       {"subunit": "Elp1", "serie": "2fold", "copies": [1], "domain": "CTD"}
                   ]
               }
           ]
       },
       "data": [
   
       ],
       "subunits": [
   
       ],
       "xlinkanalyzerVersion": "0.1"
   }
   

   Note

   Whenever you define symmetry restraints here, in the parameter file set add_symmetry_restraints = True and add 'sym_restraints' to the scoring function.