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Subsections


Modeling amino acid insertions

Here, we predict a structural model for the structurally unresolved amino acid residues 162 to 177 of chain B of 4OCM. Create the folder domain_insertion for the files generated in this section.

Structure prediction with Rosetta

For the domain insertion folding, you can take the correctly numbered full length model that was generated in section 3.1 and continue. Again, create a new folder named insertion, create a folder named full_length_model in it and copy the already generated full length PDB to it.

1
Prediction of an ensemble of possible structures:
The recommendation from the literature is to predict between 5,000 and 20,000 structures. In our test case we predict only 100 structures for demonstration purpose. We use the topology Broker as direct input to the classic Rosetta de novo protocol, wrapped by the ModelMaker package, to predict a possible ensemble of structural models. As in section 3.1, we create a configuration file fold_insertion_rpn11.tcl. Copy your configuration file with the already adapted variables to the working directory. To perform a de-novo structure prediction of the missing domain, we only need to change the analysis tasks and the commands to run Rosetta:

Execute the configuration file in VMD text mode and wait for the tasks to finish. Depending on the number of structures to generate, this may take a while.

vmd -dispdev text -e fold_rpn11_insertion.tcl

If no error occurs, go to the folder called rosetta_output_rpn11_insertion containing the results of your run.


Table 4: Rosetta domain insertion procedure arguments
\begin{table}\centering
\begin{tabularx}{16cm}{c\vert X\vert X}
arg.& descript...
... & number of structures to create & \tt\$nstruct \\
\end{tabularx} \end{table}


Interactive fitting to a cryo-EM density with iMDFF

Follow the steps explained in section 3.2.
1
Aligning the predicted model with the cryo-EM density map: See step 1 in in section 3.2.

2
Generating a density map file for MDFF:
See step 2 in in section 3.2.

3
Crop the density map:
See step 3 in in section 3.2.

4
Fitting the modeled part to the cryo EM density: See step 4 in in section 3.2.


next up previous contents
Next: Model fit to mid-resolution Up: Rosetta/MDFF Tutorial Previous: Folding protein termini using   Contents
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