tutorial-02 - Flexible-fitting of an computational RNA model into experimental data.

The prediction was downloaded from CASP15 website (https://predictioncenter.org/casp15/MODELS_PDB/R1136/R1136TS081_4.pdb). The model was docked using ChimeraX and saved as data/tutorial-02/docked.pdb. The map from the EMDB (https://www.ebi.ac.uk/emdb/EMD-14740) is in data/tutorial-02/map.mrc.gz. Secondary structure prediction using RNAView have been performed. The output is in data/tutorial-02/docked.xml.

Step 0 - Setup Conda environment

From within the tutorial directory, see if you have a conda environment set up already by running:

conda activate tempy_reff_env

If this fails, create the environment:

conda env create -f tempy_reff_env.yml -n tempy_reff_env

Step 1 - Run RibFind2

RIBFIND is used to generate heirarchical rigid body restraints. It requires the model and the RNAView secondary structure annotations. The RNAView annotations have already been generated and are saved as docked.xml. We use the --soft-cutoff for ribfind which tends to produce a deeper hierarchy which can be useful for complex fitting tasks.

ribfind \
 --model data/tutorial-02/docked.pdb \
 --rnaml data/tutorial-02/docked.xml \
 --soft-cutoff \
 --output-dir out/ribfind/tutorial-02

Step 2 - Run flexible fitting

Using the generated protein rigid bodies and the experimental map, perform flexible fitting.

tempy-reff \
 --model data/tutorial-02/docked.pdb \
 --map data/tutorial-02/map.mrc \
 --fitting-density \
 --restrain-ribfind \
 --restrain-ribfind-dir out/ribfind/tutorial-02/rna \
 --output-dir out/flex-fit/tutorial-02 \
 --resolution 8.0 \
 --platform-cuda

This will result in a director out/flex-fit/tutorial-02 containing the output of flexible fitting. The intermediate models are all stored, but the final result will be called final.pdb.

Visualisation of the model.

Step 3 - GMM refinment

Using the output of flexible fitting, you may refine the model using the more accurate GMM force field.

tempy-reff \
 --model out/flex-fit/tutorial-02/final.pdb \
 --map data/tutorial-02/map.mrc \
 --fitting-gmm \
 --output-dir out/refine/tutorial-02 \
 --platform-cuda