BASIC METHODOLOGY OF DOING HOMOLOGY MODELLING WITH ONE TEMPLATE

INPUT FILES:

1. 1LN6.pdb (template)

2. ROP.seq  (target)

3. align2d.py

4. model-single.py

5. evaluate_model.py

6. evaluate_template.py

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PART 1: ALIGNING TARGET SEQUENCE WITH TEMPLATE

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1. Convert the ROP sequence in "ROP.seq" file into PIR format and save as ROP.ali

2. Use the script in the "align2d.py" file to align the target sequence "ROP.ali" with the template structure "1LN6.pdb"

4. Go to the directory of the where modeller has been installed and paste the target file “ROp.ali”, template file “1LN6.pdb” and script files  “align2d.py”, “model-single.py”, “evaluate_model” and “evaluate_template”. MODELLER is a command-line only tool, and has no graphical user interface; instead, you must provide it with a script file containing MODELLER commands. This is an ordinary Python script. 

Run the following command

            > mod8v2 align2d.py

5. In about 10 minutes the following output files with be created

            ROP-1LN6.ali             (alignment in PIR format)

            ROP-1LN6.pap                       (alignment in PAP format)

6. Open the ROP-1LN6.pap in notepad to view the alignment conserved regions/ matching regions    can be seen with * in the third row.

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PART 2: CREATING MODEL

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1. Use model-single.py script file to create 20 models from the alignment obtained above

2. It would take about 1 hour to get the output for 20 models.

3. Run the following command from directory as before:

            > mod8v2 model-single.py

The output files will be created with the following extensions

job.log	log file
job.ini	initial conformation for optimization
job.rsr	restraints file
job.sch	VTFM schedule file
job.B9999????	PDB atom file(s) for the model(s) of the target sequence
job.V9999????	violation profiles for the model(s)
job.D9999????	progress of optimization
job.BL9999????	optional loop model(s)
job.DL9999????	progress of optimization for loop model(s)
job.IL9999????	initial structures for loop model(s)

           

           

The file ROP.B99990008.pdb can be viewed in any protein viewer.

4. Open all the model files and read the second lines of the pdb files which have the objective function value. Choose the best suitable model depending upon the minimum value of objective function and high sequence similarity. The model will the minimum objective function value / energy is the best predicted

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PART 3: EVALUATE MODEL

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1. The stereochemical properties can be evaluated using procheck

2. We can evaluate the DOPE energy of the models and compare them with templates

TARGET ENERGY PROFILE

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3. Run the following command in the directory to create the dope energy profile

            > mod8v2 evaluate-model.py

4. It will produce the output file ROP.profile

5. Open the file in notepad

6. Energy values are present in the last column of each residue row.

7. The profile can be viewed graphically using dopeProfile program (not provided in modeller, but can be seen in MATLAB, provided that it should be installed on the system.

TEMPLATE ENERGY PROFILE

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8. The energy profile for the template can also be obtained  

            > mod8v2 evaluate-model.py

9. This will produce the output file 1LN6.profile

COMPARE PROFILES

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10. To compare both profiles use MATLAB to plot the energy graphs and note the coordinates at X and Y axes of target (model) and template and analyze the results.

           

NOTE: NOTE: This procedure has been followed for all other templates as well.