Python-based
Hierarchical
ENvironment
for
Integrated
Xtallography
Contents
- cryo_fit which reports map size after step 6 before step 7 with codes of
- from iotbx import ccp4_map
- ccp4_map = ccp4_map.map_reader(user_input_map)
- target_map_data = ccp4_map.map_data()
- EMAN2's e2iminfo
- For example, e2iminfo.py <user>.mrc
- (note) As of EMAN2.2, the file extension should be mrc, not ccp4
- VMD which can visualize current map size as in this FAQ.
- For example, relion_image_handler --i <user>.mrc --new_box 370 --o <user_box_size_370>.mrc
- If a user input pdb file has unreasonable structural geometry. Although the fit between atomic model and map looks good, it is a fictitious fitting without considering ideal geometry. Run cryo_fit to find decent fit to the cryo-EM map along maintaining reasonable structural geometry.
- Providing higher (better) resolution map tends to help this problem.
- Enforcing stronger map weight tends to help this problem.
- If cryo_fit is provided a giant cryoem map with a tiny atomic model.
- Then, the cryo_fit calculates the gradient of CC because of the large empty space not filled. The constraint forces for the model are not helping as they are very small.
- 3-1. Re-run cryo_fit with an atomic model that fits the majority of the map.
- Fit multiple atomic models into a symmetric map or sequential fitting into a non-symmetric map. Watch Tom Goddard's lecture (2013)
3-2. Re-run cryo_fit with only relevant map region. A user can extract relevant map region by phenix.map_box (preferred) or phenix.map_to_model
- If the initial model is not properly aligned to a map, fit using UCSF Chimera -> Tools -> Volume Data -> Fit in Map
- Open pdb file
- [menu]
- Select -> Residue -> HIS
- Tools -> Structure Editing -> Rotamers -> OK
- (select the most probable rotamer each)
- File -> Save PDB
- Most likely, this means that initial cc is too low for MD simulation.
- When Doo Nam ran real_space_refine first, then run real_space_refined atomic model in cryo_fit, it was solved.
- Alternatively, improve initial cc by fitting initial atomic model into a map (see "How to improve initial cc?" in this FAQ)
- Less likely, but still a possible case is when the map weight is too high, lowering emweight_multiply_by may help.
"I edited out lipids, HEM and other hetero atoms and I verified that they are all gone. However, still my pdb file is not clean enough for gromacs_cryo_fit".
- Therefore, please improve initial cc by fitting initial atomic model into a map (see "How to improve initial cc?" in this FAQ)
- Doo Nam observed this error when starting molecule is not stabilized structurally. There are two possible solutions.
- Run phenix.real_space_refine before cryo_fit. Then, provide real_space_refined molecule into cryo_fit. Then, most problems were solved.
- When Doo Nam skipped minimization, this error occured. Since most users minimize the starting structure by default (cryo_fit default), most users do not need care about this. Besides, as of 9/18/2019, the default value of number_of_steps_for_minimization=20000 (which is a lot).
- It means that the map dimensions need to be larger. Therefore, check map size and solve a problem with VMD/EMAN2/relion like followings.
Like other MD simulations, gromacs need enough map box size to cover the atomic model to run (ziggle and wiggle). Refer Waters seems to be out of the box
For example, stuck-out red oxygen atoms outside the right edge of the box are the problem.
In order to run any MD simulation (including cryo_fit), a box should be large enough like
Make map box size larger (see "How to enlarge map box size?" in this FAQ), and run cryo_fit again. You can check map box size by VMD. Alternatively, remove sticking out atoms if these are unnecessary, then run cryo_fit again.
For protein modeling, I would use cryo_fit2 which is not limited by box size requirement. Most of the time, it better fits than cryo_fit1 in terms of fitting and geometry preservation anyway.
- When he provided a pdb file that has unspecified/deleted region of a molecule. Even when vmd assisted map box/cell dimension is larger than initial molecule space, this error appeared.
- When he provided a pdb file that has small number of residues (e.g. 5~16 amino acids). Even when vmd assisted map box/cell dimension is larger than initial molecule space, this error appeared. This error appeared regardless of the existence of CRYST1 and SCALE header.
- Run phenix.real_space_refine before cryo_fit. Then, provide real_space_refined molecule into cryo_fit. Then, most problems were solved.
- When Doo Nam skipped minimization, this error occured. Since most users minimize the starting structure by default (cryo_fit default), most users do not need care about this. Besides, as of 9/18/2019, the default value of number_of_steps_for_minimization=20000 (which is a lot).
- If starting molecule has non-standard chemical geometry that is too extreme that are not stabilized neither by real_space_refine nor by cryo_fit minimization, then a user needs to fix those non-standard geometry manually.
