Model-free LocScale
Reference-based local sharpening without atomic model
If no atomic model is available, or if you do not want to use prior model information, you can use the model-free mode of LocScale. This method will predict a reference map using the EMmerNet network by default and is the recommended procedure for model-free local sharpening.
Model-free LocScale workflow
Usage
locscale -hm path/to/halfmap1.mrc path/to/halfmap2.mrc -v -o model_free_locscale.mrc
Here, halfmap1.mrc and halfmap2.mrc should be the unsharpened and unfiltered half maps from your 3D refinement. If you wish to use the full map instead, use the following command:
locscale -em path/to/fullmap.mrc -mc path/to/model.pdb -v -o model_free_locscale.mrc
Point group symmetry
If your map has point group symmetry, you need to specify the symmetry to force a symmetrised reference map for scaling. You can do
this by specifying the required point group symmetry using the -sym/--symmetry flag, e.g. for D2:
locscale -hm path/to/halfmap1.mrc path/to/halfmap2.mrc -v -sym D2 -o model_free_locscale.mrc
The output will be a locally sharpened map scaled according to the scale factors derived from the EMmerNet-predicted reference map.
Recommended use of unfiltered input maps
Note that using unfiltered maps as input is essential. If using previously filtered maps, information beyond the spatial filter cutoff cannot be recovered.
Model-free local sharpening with pseudomodels
Another option for model-free sharpening is use a full pseudotatom model. This can be enabled by passing the --build_using_pseudomodel flag when invoking LocScale. This mode will estimate the molecular volume using statistical thresholding and generate a pseudo-atomic model within the thresholded boundary to approximate the distribution of atomic scatterers and estimate the local B-factor. It will then generate an average reference profile for local sharpening based on the experimental data and expected properties for electron scattering of biological macromolecules [2]. Use this if the default EMmerNet-based reference map generation does not work well for your data (e.g. if the map is too noisy or if the map has very low resolution).
locscale -hm path/to/halfmap1.mrc path/to/halfmap2.mrc -v -o model_free_locscale.mrc --build_using_pseudomodel
Usually all default parameters for pseudomodel and reference profile generation are fine and we do not recommend to change them, but you can change them if you deem fit.
Point group symmetry
If your map has point group symmetry, you need to specify the symmetry to force the pseudomodel generator for produce a symmetrised
reference map for scaling. You can do this by specifying the required point group symmetry using the -sym/--symmetry flag, e.g.
for D2:
locscale -hm path/to/halfmap1.mrc path/to/halfmap2.mrc -v -sym D2 -o model_free_locscale.mrc
The output will be a locally sharpened map scaled according to the local scale factors derived from the ADP distribution of the hybrid pseudoatom model.
Speed-up computation on multiple CPUs
To speed up computation, you can use multiple CPUs if available. LocScale uses OpenMPI/mpi4py for parallelisation, which should have been automatically set up during installation. You
can run it as follows:
mpirun -np 4 locscale -hm path/to/halfmap1.mrc path/to/halfmap2.mrc -v -o model_free_locscale.mrc -mpi
ge of multiple CPU cores by using joblib. In this case, simply specify the num
ber of CPU cores using the -np flag as follows:
```bash
locscale -hm path/to/halfmap1.mrc path/to/halfmap2.mrc -np 4 -v -o model_free_locscale.mrc
```