Team:Goettingen/notebook drylab

Homology modelling procedure

To generate our models we followed the procedure from the basic and advanced tutorials of the Modeller web page with some modifications. We first selected ten templates with known structures for our peptide-contaning scaffold. In the case of the protein G scaffold, this was done using the best five results from the Modeller "profile.build()" command and the best five results from an homology search in the Protein Data Bank; the PDB files of the selected templates are: 1PGB, 1FD6, 1FCL, 1QKZ, 1EM7, 2GB1, 1GB1, 2RPV, 2QMT and 3GB1.

In the case of the GFP scaffold, the selected templates were only those from the results of the "profile.build()" command; their PDB files are: 1XSS, 1OXD, 1XA9, 1MOU, 1XMZ, 1UIS and 1GGX.

Before starting with the alignments, we made a secondary structure prediction using the GeneSilico Metaserver. The consensus data was parsed from the results of the metaserver and processed so that the secondary structure information was associated with their corresponding amino acid residues.

The PDB files from the selected templates were aligned using the "salign()" command to generate a template alignment and then the sequence of each of our peptides was aligned separately to the template alignment using the ".append" method. Ten initial models were generated for each peptide using the secondary structure data as restrains; the best model with the best DOPE score was selected for loop refinement. The loop refinement generated ten models for each peptide; this was iterated until no improvements were observed in the DOPE score of the resulting structures.

References

• 1. Kurowski MA, Bujnicki JM. GeneSilico protein structure prediction meta-server. Nucleic Acids Res 2003; 31: 3305-3307



• 2. N. Eswar, M. A. Marti-Renom, B. Webb, M. S. Madhusudhan, D. Eramian, M. Shen, U. Pieper, A. Sali. Comparative Protein Structure Modeling With MODELLER. Current Protocols in Bioinformatics, John Wiley & Sons, Inc., Supplement 15, 5.6.1-5.6.30, 2006.




• 3. M.A. Marti-Renom, A. Stuart, A. Fiser, R. Sánchez, F. Melo, A. Sali. Comparative protein structure modeling of genes and genomes. Annu. Rev. Biophys. Biomol. Struct. 29, 291-325, 2000.




• 4. A. Sali & T.L. Blundell. Comparative protein modelling by satisfaction of spatial restraints. J. Mol. Biol. 234, 779-815, 1993.




• 5. A. Fiser, R.K. Do, & A. Sali. Modeling of loops in protein structures, Protein Science 9. 1753-1773, 2000.