Tutorial:Structure Visualization with Chimera and Cytoscape

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Slideshow Structure_Visualization_with_Chimera_and_Cytoscape (about ?? minutes)
Handout StructureViz.pdf (?? pages)

Tutorial Sources
Tutorial Curators Anna Kuchinsky, Scooter Morris

Structure Visualization with Chimera and Cytoscape

Biological Use Case: Identify protein-protein interactions at a deep level by examining structural information. In this case, we will look at breast cancer data.

Getting Started

  1. Install Cytoscape
  2. Install the structureViz plugin.
    • Go to Manage plugins->Available for Install->Analysis->structureViz.
  3. Install UCSF Chimera, by going to http://www.rbvi.ucsf.edu/chimera/download.html.

Load Data

You can load the sample data pte.xgmml directly onto Cytoscape...

  1. Go to File->Import->Network (multiple file types)...
  2. In the dialog box, select Remote for Data Source Type and enter: http://www.rbvi.ucsf.edu/Research/cytoscape/structureViz/pte.xgmml into the data field.
  3. Press Import.

You can also import the sample data as a file...

Importing a Network File
  1. Download pte.xgmml and save it to your disk.
  2. Launch Cytoscape.
  3. Go to File->Import->Network (multiple file types)...
  4. When the dialog box pops up, select Local for Data Source Type and press Select.
  5. This will bring up a file browser. Browse to where you saved pte.xgmml and select it.
  6. Press Import.

The network loaded contains the two subgroups of the amidohydrolase superfamily.

  • Nodes represent the proteins in the two subgroups
  • Edges represent the similarity between the proteins
    • Edges contain information about the E-values calculated using the UCSF Shotgun program.
  • The network was laid out using the Cytoscape edge-weighted, spring-embedded layout algorithm:
    • Layout->Cytoscape Layouts->Edge-weighted->Edge-weighted Spring Embedded->BlastEValue

Initial Test

Once you have successfully loaded the file, Cytoscape should look like this.

Phosphotriesterases Loaded Network

The next step is to see if structureViz is loaded correctly and test connectivity to Chimera.

  1. Select a number of nodes in the Cytoscape window by dragging your mouse over the nodes. The nodes you select should turn yellow.
  2. Get a list of the structures by going to Plugins->Sequence/Structure Tools->Open Structure(s). This should display a list of all structures.
    • If the Sequence/Structure Tools menu isn't available, structureViz was not correctly installed.
    • If no PDB structure IDs are shown in the list, then the network was not correctly loaded.
  3. If everything works, select the structure 1EZ2, which is one of the structures for phosphotriesterase, GI 2098312.
    • If you get an error dialog, it probably means that the Chimera application is not on the default path. This is common for MacOS X, but could also happen in Linux and Unix installations.

Configuring Chimera's Path

Cytoscape Preferences Editor

If you did not get an error dialog box, skip this step. If you got the error dialog, you need to configure the path to Chimera. You need to add a new property: structureViz.chimeraPath using the Cytoscape Preferences Editor.

  • Go to Edit->Preferences->Properties. This will bring up the Preferences Editor.
  • Click Add and enter the name of the property: structureViz.chimeraPath.
  • Click OK and enter the path to your Chimera application.
    • On a Mac, this should be the path to the Chimera executable, not the Chimera.app directory.
    • If you install Chimera in your Applications folder (the recommended install location), then set the path to /Applications/Chimera.app/Contents/MacOS/.
  • Once you have set the path, click OK.
  • Select the checkbox next to Make Current Cytoscape Properties Default. Click OK again. The program will now remember these settings.

Browsing and manipulating structures

Cytoscape window with Molecular Structure Navigator and UCSF Chimera

Several things should have happened when you requested structureViz to load 1EZ2.

  1. Chimera should launch
    • go to the RCSB website and load the coordinate information for 1EZ2
    • then display the model.
  2. The Cytoscape Molecular Structure Navigator should pop up.

You can manipulate the structure by

  • using the menus provided by the Cytoscape Molecular Structure Navigator
  • using the menus and commands provided by Chimera directly.
    • to move and scale the structure with your mouse, you will need to interact directly with the Chimera window.

Using the Cytoscape Molecular Structure Navigator

The Cytoscape Molecular Structure Navigator provides a tree-oriented interface (similar to the Windows file browser or the Macintosh finder). At the top level, all of the models currently loaded are on display. One level down, within each model is the list of chains, and finally, within each chain is the list of residues.

First, we'll explore the structure of 1EZ2.

  • You can see that 1EZ2 has two chains defined, Chain A, and Chain B.
    • Each chain includes the "het" atoms and the solvent that is associated with the structure.
  • Open up the Chain A
    • Scroll down until you see ZN401 and ZN402
      • they are before any of the solvent molecules: HOHxxxx
    • Select one of these
      • The selected name highlights, as you would expect
      • The ion will be outlined in green in the Chimera window
      • The node that refers to the 1EZ2 structure (gi2084365) will be highlighted in green in the Cytoscape window.

An easier way to select all of the "het" atoms is to use the Molecular Structure Navigator Select menu.

  • Use the menu to do a Select->Ions
    • All of the zinc (ZN) atoms selected.
  • Now do a Select->Ligand
    • The diisopropylmethyl (DII) ligands that are bound to the structure will be selected.
    • NOTE: Many structures will have ligands defined, and when they are defined, they are not always segregated out into a separate chain. PDB structure definitions are quite varied so some exploration of the structure through the Molecular Structure Navigator is usually warranted.
  • Take a look at "Chain A".
    • Open it up and notice that by default, the residue names use the standard three letter abbreviation.
    • The View menu allows you to set either the full name View->Residues as->full name...
      • (for those of us who constantly confuse asparagine with aspartic acid)
    • ... or the single letter abbreviation View->Residues as->single letter'
    • When the display parameter is changed, the tree will need to reload itself so it will return to the initial closed state.

Using Chimera

  • Details on using Chimera's command or menu interface is beyond the scope of this tutorial.
  • Users interested in exploring the features provided directly by Chimera are encouraged to review the Chimera User's Guide.
  • The table below describes the default mouse manipulation commands.
  • Detailed information on mouse manipulation is available in the Mouse Manipulation of Models section of the Chimera User's Guide.
    • Note that the table below assumes a three-button mouse.
      • On a Mac with a one-button mouse, buttons 2 and 3 can be emulated with the option and apple (clover) keys, respectively.
Mouse Manipulation of Models in Chimera
Manipulation Mouse Cursor
XY Rotation Button 1 (left button) within the central regino of the graphics window Small circle
Z rotation Button 1 (left button) in the periphery of the graphics window Two curved arrows
XY translation Button 2 (middle button) Cross formed by two double-headed arrows
scaling Button 3 (right button) Diagonal double-headed arrow enclosing a small square
Z translation Ctrl-Button 2 (control key + middle button) Vertical double-headed arrow

Enhance the display

As we noted before, the structure 1EZ2 includes two polypeptide chains, numerous water molecules, and two ligands. We want to simplify the structure to improve the visualization of the catalytic pocket as well as prepare the structure for doing alignments.

Delete unnecessary residues

We will start by deleting all the residues that are not necessarily for either the visualization of the pocket or the alignment.

  • Open up Node gi12084365 [...] so that all of the chains are visible.
  • Select all of the water molecules by doing a Select->Solvent
  • Bring up the generic pop-up menu (Button 3 in an empty area of the dialog)
    • Choose Delete selection to delete all of the waters in the structure.
      • You will be asked to confirm by clicking Yes and then the menu will collapse.
  • The waters will be deleted from the Chimera structure as well as from the Molecular Navigator.
    • This will also cause the Node menu to collapse.
  • Open the menu back up
    • Delete Chain B by bringing up it's pop-up menu (Button 3 again)
      • Choose Delete selection.
Cytoscape after deletion of unnecessary residues

Simplify the chain and change the depiction

You should now be left with a structure in the Chimera window that includes only chain A. Next, we will use the Cytoscape Molecular Structure Navigator to simplify the display and allow us to focus in on the functional residues.

  • Make sure Chain A is still selected and bring up the popup menu again by pressing Button 3 or Ctrl+Button 1.
  • Select Show->Backbone only.
    • This will remove all of the side chains and display only the protein backbone.
  • Now, change the depiction of the molecule to a ribbon.
    • Select Chain A, bring up the popup menu, and select Depict->Ribbon->Round.
      • This will render the protein backbone as a ribbon.
  • To color the ribbon, press Color->Rainbow by residue.
  • Clear the selection by using the Select->Clear selection menu in the top menu bar.

Your Chimera screen should look like this.

Ribbon Backbone of Chain A of 1EZ2

Visualize the functional residues and ions

The final step in visualizing our structure is to view the functional residues and ions in the catalytic pocket. This will utilize many of the same functions we saw before, plus the ability of structureViz to use the Functional Residues attribute associated with the nodes in Cytoscape.

Select the functional residues

  1. Select the Node and bring up the popup menu.
  2. Select Select->Functional Residues. You will see sections of the ribbon highlighted in green. The Molecular Structure Navigator will also open up to display the selected residues.

Display the color and functional residues

Now that the functional residues are selected, we can display their side chains and change the color (we could change their depictions in other ways).

  1. Position your mouse in the Molecular Structure Navigator into the blank space to the right of the displayed residues.
  2. Bring up the popup menu. The actions in this menu will apply to all selected residues (or chains, models, etc.)
  3. Now, press Show->All to display the side chains of the functional residues.
  4. Go to Color->Residues->magenta to change the color of the side chains so that they will stand out a little.

Select, display, and color the ions

Chimera's ability to manipulate molecular structures by chemistry is reflected in the features of the Molecular Structure Navigator. To select the ions, we can either use the popup menu of the Node level of the structure to select only the ions of that structure, or the top menu bar Select menu to select all ions. In this demo, we'll use the Node approach.

  • Bring up the Node gi12084365 [...] popup menu.
  • Go to Select->Ions.
  • Show the ions by bringing up the background popup menu (the one in the blank space to the right of the displayed residues).
  • Press Show->All to display them
    • SelectColor->Residues->yellow
    • SelectDepict->Sphere to make them stand out.
      • If you move the structure around in Chimera, you should be able to focus in on the catalytic pocket and get an image similar to this.
Catalytic pocket of 1EZ2

Comparing Structures with structureViz and Chimera

The network pte.xgmml has two SFLD subgroups: the phosphotriesterase like group and an unknown group labeled as unknown119. Proteins in these groups are very similar, but only the proteins in the phosphotriesterase like subgroup are known to hydrolyze organophosphates. One way to investigate possible reasons for the differences in catalytic function between these two very similar groups of proteins is to compare them structurally. Using structureViz and Chimera you can load multiple proteins into Chimera and align them. In our case, we want to align 1EZ2, the structure for a phosphotriesterase from Brevundimonas diminuta with 1BF6, the structure for one of the unknown proteins from Escherichia coli.

Add 1BF6 to the Visualization

  • Back in Cytoscape, bring up the pop-up menu for node gi5542102 by placing your mouse over the node and doing a right-click (Button 3).
  • Now load the structure by doing a Structure Visualization->Open structure(s)->1BF6.
    • This will load 1BF6 into the same Chimera session as 1EZ2.
  • Clean up the structure by deleting Chain B from 1BF6 and all of the waters.
  • You may also want to depict the remaining structure as a ribbon.

Align 1EZ2 and 1BF6

We now have two structures, 1EZ2 and 1BF6. What we would like to do is look at a structural comparison between the two structures to determine if it gives us any clues about the functional differences. We'll do this using Cytoscape Molecular Structure Navigator's Align Structures menu and Chimera's MatchMaker tool. MatchMaker provides a protein superposition by first constructing a sequence alignment and then performing a least-squares fit to superimposed the aligned residue pairs.

  • Under the Chimera menu, select Align Structures->by model
    • (we could have done an align by chain, but we've already limited the structures to a single chain, so that isn't necessary).
    • This will bring up the Cytoscape/Chimera Structure Alignment Dialog.
  • In the menu within the Reference Structure box, select Node gi12084365; model 1EZ2.
  • In the table underneath, select 1BF6, then click on the Align button.
    • This will perform the alignment using Chimera's MatchMaker tool.
    • After the alignment is complete, the Cytoscape/Chimera Structure Alignment Dialog will display the results, including:
      • the number of aligned pairs of residues,
      • the RMSD (root mean squared distance) between the pairs, and
      • an overall score that takes both of those into account.
    • In this case, the RMSD is 0.992, which indicates a very close alignment, at least amongst the 214 aligned pairs.
  • Look at the resulting superposition to see what differences might be visible. In particular, note the similarity of the catalytic pocket, but also note the entrance to the pocket is very different between the two structures.
Alignment of 1EZ2 and 1BF6

Exiting structureViz

Once you have completed viewing, comparing, or manipulating your structures, you can close the Cytoscape Molecular Structure Navigator (and Chimera) by selecting Chimera->Exit. This will close the dialog and Chimera and you will be back in Cytoscape.