Task homologyModelling 2011

For the sequences used in this practical, protein structures have been determined. However, in real-life projects, you often do not have structures. Therefore, we will use structure prediction methods to predict the 3D structures of our sequences. We will also check whether and how the SNPs change the predicted structures.

Theoretical background talks

We will be looking at two parts of homology modelling (-> two talks)

• Identifying suitable templates and producing and alignment
• Calculating the actual models and evaluating the results

Please include these programs in your talks

• HHpred
• Swissmodel
• Modeller
• iTasser

The talks should cover

• how the methods work behind the scenes,
• some information on their performance, strengths and weaknesses (as e.g. seen in CASP),
• a brief intro into how to call them and where to find documentation,
• some discussion how to evaluate the resulting models

Calculation of models

• Get an overview of available homologous structures based on the sequence searches and alignments.
  • Use HHsearch to check whether you can extend your list towards more remotely similar structures
• Divide your homologous structures into groups at
  • > 60% sequence identity
  • > 40% sequence identity
  • < 40% sequence identity (ideally go towards 20%)
• If possible (i.e. if there are structures at that level of sequence identity) create models with Modeller (command line), Swissmodel and iTasser (online) using one template from each of the groups.
  • Use the default settings of the methods, i.e. use the standard workflow and directly feed the alignments to the modelling step
  • In addition: Have a look at the alignments you use for modelling.
    • Collect sequence-based information (important residues, sequence family profiles, secondary structure prediction, etc.) to check the alignment.
    • Edit the alignment.
    • Then, proceed with modelling.
    • Document what you changed and why.
  • In addition: For modelling with Modeller: Use more than one template in one modelling step (if possible due to availability of templates).
• Feed your (subjectively best) models of each category into 3D-Jigsaw to get out recombined, optimised (?) models.

Now, you should have quite a large number of models.

Evaluate your models

• Check the numeric evaluation of your models (scores given by the modelling programs)
• Compare the models to the experimental structure (Select one apo and one complex structure if there are several experimental structures, document your choice of reference)
  • Calculate the TM score of the models.
  • Calculate the C_alpha RMSD of the models.
  • Extra diligence task: define a radius of 6 Angstrom around the catalytic centre and calculate the all atom RMSD in that region
• Discuss your results:
  • Is any method systematically better at predicting the structure?
  • Does this depend on the similarity of the template?
  • Does refining the models by hand help?