Building 3D Structural Alignments



Introduction

CE and FATCAT are two pieces of software that compare and align protein 3D structures in PDB format and can be accessed via a web interface.

In their results, they both provide
In our teaching, we are interested in examining structural alignments, not with a view to understanding the methods and algorithms used to build them - rather, as a tool to understanding the interpretation of sequences alignments in a structural context. Thus, below we provide only a simple description of how to provide appropriate input to the software, and how to visualise the resulting alignment in 3D.

Obtaining input PDB files

Both servers can calculate alignments when provided with a pair of PDB format 3D protein structure files.

There are many different ways to identify files for the set of proteins you might be interested in e.g.
The links to the PDB files are sometimes tricky to find - the image below shows the mouse pointer positioned over the icon that leads to the PDB format file for the record being downloaded.



Note that one PDB file may contain the structures of more than one protein. The different polymers in such a file are specified by their "chain" identifiers - for example, in the PDB file 2w8l, there are three polymer chains
If you are comparing and aligning a pair of protein structures, it is therefore important that you know which chain within the PDB file corresponds to the protein you want to include in the alignment.

You can determine which proteins correspond to which chains by examining the PDB file directly, or usually much easier, examining the corresponding pages describing that particular PDB file at PDB sum or PDB at the RCSB. The links below lead to the corresponding records for 2w8l from these two resources

Example files

These two PDB files are both bacterial adhesins

Viewing aligned structures in PyMOL

To highlight the superposition of the aligned structures in the resulting PyMOL files, try typing the set of commands given below into the typed-command interface of the PyMOL window. This will display the two structures in a similar way as seen in the image below. This colouring and representation makes a good starting point to compare the aligned regions of the chains.
Finally, use the instructions on this page PyMOL to switch on the single-letter amino acid cod sequence view at the top of the window.

CE Webserver Pairwise Structural Alignment

Begin with the "CE calculate two chains" structural alignment query page.

For each of the two chains:
Then click the "Calculate Alignment" button near the top of the page to begin calculating the alignment.

For example, before submitting the two example structures for alignment, the query form would look similar to the image shown below:


The result page shows the pairwise sequence alignment corresponding to the calculated pairwise structural alignment.

To obtain the file with the two aligned structures in PDB format, follow the link "Download alignment as a PDB file" below the sequence alignment.

NOTE - certain problems associated with loading the aligned structure files into PyMOL are avoided by making sure that the file is saved with the suffix .pdf i.e. it is strongly recommended that you name the file(s) you work with something like "ce_4hhb_a_vs_4hhb_b.pdb" rather than "ce_4hhb_a_vs_4hhb_b.txt" or similar!

This is the CE pairwise structural alignment PDB file for the two example structures.

This link shows the result page for the two example structures.

To view the 3D alignment PDB format file in PyMOL as shown above, you will need to open the file in a text editor, and delete the following two lines that are found almost in the middle of the file, re-saving the file under a different name:

ENDMDL
MODEL 2

This link is to the CE PDB alignment file for the two example structures modified in this way.

Alternatively, open the un-modified alignment file in PyMOL and use the menu to select:

Movie->Show all states

This will also show the two structures at the same time, although without the cartoon secondary structure representations.

FATCAT Webserver Pairwise Structural Alignment

Beginning with the FATCAT pairwise alignment query form, for each of the two structures:
And example of how the form will look at submission is shown below

Once the results page is returned, follow the link "View the alignment file" on line "2" of the results to see the pairwise sequence alignment corresponding to the calculated pairwise structural alignment - this link is to the corresponding sequence alignment for the two example bacterial adhesin structures

The pairwise structural alignment can be viewed following the link from the second bullet point of the "4" results "Download the pdb file" - this link is to the alignment calculated for the two example bacterial adhesin structures.

This link shows the results page for the two example structures.

The pairwise structure can be loaded directly into PyMOL.

Author: Aidan Budd
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