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 Protein Homology/analogY Recognition Engine V 2.0


 

Help



Phyre Investigator

Phyre Investigator is a 'workbench' for analysing a range of features of your protein sequence and a Phyre2 model of your protein.
Please be aware that Phyre Investigator is in Beta-testing. I am very keen to hear feedback from users regarding problems they have with the interface, web browser compatibility, strange results, or any suggestions on how to improve the system. Please contact me (Harry Powell), with any comments or problems.
 
Your detailed template results will now look something like this:
Clicking on the 'Run Investigator' button will let you perform the analyses on this model of your sequence. The green tick and link takes you to results of a previously submitted Investigator run.
 
Below is an example of the interface:
 
Currently, the following analyses are performed:
  • Model quality assessment by ProQ2
  • Alignment confidence from HHsearch
  • Clashes, Rotamers, Ramachandran analysis by Molprobity
  • Disorder prediction by Disopred
  • Pocket detection by fpocket2
  • Catalytic site detection from the CSA
  • Mutational analysis by SuSPect
  • Conservation analysis using Jensen-Shannon Divergence
  • Interface detection using the protindb and PI-site databases
  • Detection of other features using the Conserved Domain Database

Navigating the interface

The screen is divided into 3 main sections from top to bottom. The information box, the structure view and analyses buttons and the sequence view
The structure view and analyses section is divided into 3 sections, from left to right: The The JSmol interactive viewer, the Analyses buttons, and two graphs showing sequence profile and mutational predictions.
 

Analysis buttons

Clicking on an analysis button will display, in the information box, a brief summary of whichever analysis is currently active and links to downloadable raw data. It will also colour the structure in the JSMol view in accordance with the analysis chosen and a colour-coded key to the left of the structure. Finally it will add an extra row to the sequence view, illustrating the same information but in a sequence context.
 

Sequence View

The sequence view displays the predicted secondary structure of your sequence, the confidence in this prediction, the secondary structure of the model, your sequence and which regions have been modelled. In addition, once you've clicked on an analysis button, an extra row is added showing the corresponding information from the analysis in a sequence context.
Hovering over a sequence position will highlight that position with vertical bars to either side of the residue in question. It will also highlight that residue in the JSMol 3D viewer as a red halo around the atoms of that residue. Finally, it will also show the appropriate sequence profile and mutation graphs for that position.
Clicking on a residue will cause that residue to be spacefilled in the JSMol viewer. You may select multiple residue by repeated clicking. At any time you can clear your selection by clicking the "Clear selection" button above the sequence view. You may also take a snapshot of the structure at any time using the "Take JMol snaphot" button.
 

Sequence Profile Graph

The sequence profile graph represents residue preferences in your protein at a particular sequence position. These values are calculated by scanning your sequence against a large sequence database using the iterative searching program PSI-Blast.
In the example to the left, the 20 possible amino acid types are labelled along the x-axis with their one-letter code. The coloured bars indicate the favourability of each residue type at the currently highlighted position in your sequence. In this particular position in the sequence, we can see that I and L are highly favourable (tall and yellow), M and V are moderately favourable (quite tall and green) whilst the remaining amino acids are generally unfavourable. In particular G is the most unfavourable at this position.

Mutational Analysis Graph

The mutational analysis graph represents the predicted effect of mutations at a particular position in your sequence. These predictions are made using the SuSPect method, described below.
In the example to the left, the 20 possible amino acid types are labelled along the x-axis with their one-letter code. The coloured bars indicate the probability that a mutation to the corresponding residue will have some effect on function of the protein or on the phenotype of the organism. In this particular position in the sequence, we can see that many mutations are likely to effect function, the highest likelihood coming from mutations to D, G or P. However, Y is very low suggesting it is the wild-type. However, there are several important caveats.

The SuSPect method

The SuSPect method is available as a standalone web server here, where you will find more information about how the method works and more options for uploading sets of sequences, viewing pre-calculated results for the entire human proteome and more.
When using SuSPect through Phyre Investigator, it is important that your sequence is the wild-type. Submitting a mutant protein to Phyre2 and then Investigator will lead to misleading predictions from SuSPect.

Citing SuSPect: SuSPect: Enhanced Prediction of Single Amino Acid Variant (SAV) Phenotype Using Network Features
Yates et al. J Mol Biol. 426(14): 2692-2701 (2014)
 

Future

More help and a video tutorial for Phyre Investigator will be available in the coming weeks. I hope you find the new system useful in your research.
Harry Powell


Phyre is now FREE for commercial users!

All images and data generated by Phyre2 are free to use in any publication with acknowledgement

Accessibility Statement
Please cite: The Phyre2 web portal for protein modeling, prediction and analysis
Kelley LA et al. Nature Protocols 10, 845-858 (2015) [paper] [Citation link]
 
© Structural Bioinformatics Group, Imperial College, London
Michael Sternberg 
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