Pymol Practice

Published

August 21, 2023

Modified

February 7, 2025

1 Note about installing Pymol

There are two options to use Pymol in your computer:

  1. Edu license:

  2. Open Access version from GitHub:

    • On Linux: https://pymolwiki.org/index.php/Linux_Install

    • On MacOs: https://pymolwiki.org/index.php/MAC_Install

    • UPDATE: for new MacOS you may need to install some dependencies:

      brew install tcl-tk

  3. Open Access version from conda-forge channel:

    conda install -c conda-forge pymol-open-source

2 Part A: A 10-steps self-guided practice to learn Pymol

The first thing to do is to open PyMOL. You can do that by simply double-clicking on the icon or typing pymol in a Terminal window. From here and on, we will just use PyMOL by typing in the command line on the small window on top where it says ‘PyMOL>’. Feel free to explore the graphical interface on your own.

The next thing we are going to do is navigate to the directory (folder) where we want our files to be located. By default (in Linux and MacOS), Pymol opens in your home directory. Inside PyMOL, we can use Linux commands to navigate to that directory. If you type ls, you will see the files and folders that exist in the home directory and if you type the cd command and providing a path you can chance your working directory.

2.1 1. Get a structure.

As an example, we are going to work with the PDB structure 6KI3. There are two ways to get a structure ready to work on Pymol, you can open a file in your local machine or you can fetch it from the PDB.

  • If you have already the PDB go to File menu > Open PDB

  • You can also fetch the directly from the PDB database, either by menu File > Get PDB and typing our PDB ID or by Command line:

fetch 6KI3
Note

Note: You can customize the download by specifying one chain or adding the electron density map in the popup window or by command line. Check the wiki: https://www.pymolwiki.org/index.php/Fetch

Note2: If you want to download structural models from AlphaFold you can install Alphafold2import plugin from here: https://github.com/APAJanssen/Alphafold2import

What do you see?

First thing you should do is playing around with your mouse. Most of you will have a 3-Button mouse. The panel in the right-bottom corner contains a description of mouse controls under the Viewing mode. You can also check it here. If you have a 2-Button mouse, you can find more information about how to control the mouse here.

Now, try to describe the structure components.

What are the small red dots and the big spheres?

Crystallography structures show protein oligomers and may also contain, enzyme substrates, cofactor or solvent molecules (water and ions). For most purposes, we may prefer to work only with a monomer and remove the water molecules.

There are two options to remove the water molecules:

  1. Use the Hide menu on the right hand of the screen (marked with the yellow arrow in the picture above).

    • H>waters

  2. On command-line you can also hide custom solvent:

hide everything, solvent
show wire, solvent #to bring it back
show nb_spheres, solvent #alternative display
  1. Use the Show menu
  • S>wire>not_bounded

As you can imagine, there are many different ways of visualizing the molecules and features you can show on the screen that you can explore on your own. In pymol we have lines, ribbon, surface, everything, mesh, volume, cartoon, spheres, labels and sticks.

Like in the example above, you can hide/show any representation for an object or selection using the following sintax:

hide \[representation\], \[selection-name\]
show \[representation\], \[selection-name\]

The selection-nameis an optional argument and can be very complex or specific, involving one or more portions of one or more objects.    

Tip

❗️You can see a detail explanation of the Pymol selection syntax in the Pymol wiki here.

Starting in version 1.5, PyMOL contains an undo command (also Ctrl+Z or Cmd+Z). However, it only affect object edition (changes in the structure), no changes in visualization or diplaying features. If you made a mess in the visualization of your molecule(s), you only have two alternatives.

  • Use File > Reinitialize to reset PyMOL to its initial state, but all work will be lost.

  • Save your session when you accomplish a wanted display:

    • A PyMOL sessions retains the state of your PyMOL instance. You can save the session to disk and reload it later. You can setup a complicated scene, with transitions and more, and simply save it as a PyMOL Session (.pse) file. Use File=>Save Session or Save Session As…. or the command save https://pymolwiki.org/index.php/Save

    • Loading sessions is just as easy: File=>Load and choose your session file with all your molecules with the same visualization.

    • You can also save temporal scenes within your session, like frozen screenshoots while keep working, using the command scene: https://pymolwiki.org/index.php/Scene

2.2 2. Choose representation and color schemes.

Different types of representation can be selected for each object or selection with the button S on the right panel. There are also wide range of flavors for each representation. For example, for cartoons: https://pymolwiki.org/index.php/Cartoon. For instance, we can color the protein by secondary structure using Menu C

Spectrum color scheme is also useful to follow the secondary structure succession.

Using upper menu (Settings and Display) we can also change several features of the image. Try yourself!

It’s faster if you just type:

color tv_orange, ss h
color paleyellow, ss s
color grey, ss l+
set sphere_color, tv_red
bg_color white

Note: l+ will color loops and other unassigned residues, including DNA

🔝🔝Check other coloring alternatives: https://pymolwiki.org/index.php/Advanced_Coloring

For DNA: https://pymolwiki.org/index.php/Examples_of_nucleic_acid_cartoons

Example:

set cartoon_ring_mode, 3
set cartoon_nucleic_acid_mode, 4
cartoon oval
cartoon automatic, 6KI3 and chain A or chain B
set cartoon_ring_width, 0.2
set cartoon_ring_color, deepsalmon
set cartoon_nucleic_acid_color, deepsalmon

2.3 3. Create new object and/or get rid of what you don’t want

If we want to work only with some objects or chains in the PDB file, you can do it in several ways. There are several options but you need to show the sequence (Menu Display or button S on the botton-right corner).

You can remove objects (Option 1A or 1B) or copy a selection to a new object (Option 2A or 2B). You can test all with the instructions below, but I find more convenient to use command line alternatives (1B and 2B).

⚠️️ Note that the options 1 and 2 below are alternative ways to get the same results. Try only one option each time starting over from 6KI3 structure.⚠️❗️

  • Option 1A: select in the sequence panel the sequence you want to remove and remove it

    remove sele

  • Option 1B: use command line

    • Step 1: First you need to see the composition of your structure
    get_chains 6KI3
select chain A
select chain B
#to see who is who
    • Step 2: select the chains you want to remove and remove them
    select kk, chain A or chain C or chain D or chain E
delete kk

  • Option 2A: Select in the sequence panel the sequence or whole chains you want to keep: A > “Copy to object” or “extract to object” will allow to work independently with that portion without removing the rest. Just click on the object to disable.

  • Option 2B: use command line

    • Step 1: Select and create:
    select kk, chain B or chain F or chain G or chain H
create monomer, kk

Or directly create an object with your selected chains:

create monomer, chain B or chain F or chain G or chain H

  • Step 2: Now you can remove the original object (if you wish)

    remove 6KI3

Some interesting points about selection on Pymol:

Create your own representation of the monomer ternary complex (protein + metal cofactor + DNA)

2.4 4. Compare structures

Download some related structures, like 6KHY, 3AAM and 2NQH.

Note that 6KHY also contains several chains. For comparison we don’t need the DNA, so we will use only the protein chain A.

fetch 6KHY
create 6KHY_monomer, 6KHY and chain A #it also has several chains
delete 6KHY #remove the original structure for clarity
fetch 3AAM
fetch 2NQH

Now you should have four structures: 6KI3 (or monomer), 6KHY_monomer, 3AAM, and 2NQH

Pymol has several alternatives for structural alignments. The three more common are the commands super, alignand cealign, depending on the level of similarity. super is very fast and competent when proteins sequences are very similar, whereas align is a very quick alternative and useful for most purposes with a clear sequence similarity. On the other hand, cealing is based in the CE algorithm and although somewhat slower it provides better structural alignment for proteins without clear similarity.

align 6KHY_monomer, 6KI3
align 3AAM, 6KI3
align 2NQH, 6KI3
zoom all #to center the view of all the proteins in the new position

Can you range the three new structures by their similarity with 6KI3?

Multiple superposition

From version 1.7.2, there is also a command to align multiple structures with a reference molecule, using extra_fit.

2.5 5. Find what you are looking for

Now we can play with different representations for specific residues, like catalytic ones: D231, H182, H115, E142, H218, H78, E271, H233.

You can select them directly picking from the sequence. If you want to keep this selection for the future you can name it in the A menu. Using show and color menu, you can also change how those residues are displayed.

Quick alternative (note the different use of and-or and the parenthesis https://pymolwiki.org/index.php/Selection_Algebra):

select catalytic_res, monomer and (resi 231 or resi 182 or resi 115 or resi 142 or resi 218 or resi 78 or resi 179 or resi 271 or resi 233)
show sticks, catalytic_res
zoom catalytic_res

There are also some interesting tricks for disclosing ligand binding or catalytic sites:

More info about selection of multiple objects in Pymol here: https://pymolwiki.org/index.php/Selection_Algebra

2.6 6. Highlight what you want

You can also hide the cartoon partly or totally and color by elements

hide cartoon, monomer and chain A
show sticks, chain D and resi 10
show sticks, catalytic_res
set_bond stick_radius, 0.25, catalytic_res
color red, chain D and resi 10 and elem o

2.7 7. Show labels

  • Use the L menu to show the labels of your object/selection

  • You can also use the command label to show labels

label [selection], [expression]

Labels can be formatted in different ways, for example:

set label_outline_color, black
set label_font_id, 10
set label_size, -2.5
set label_position,(0,0,100), resi 87

You can also move the labels with “Edit mode”: http://polo.engr.colostate.edu/drillbits/lessons/59/#:~:text=To%20get%20labels%20exactly%20where,click%20to%20move%20the%20label.

Be careful not to pick on the molecule (UNDO still partial!)

Make a picture of the catalytic center with your custom representation, including labels

2.8 8. Measure distances

Menu Wizard>Measurements

Click “Done” on the right when you are done.

2.9 9. How to make a high-res picture: RAY IT!

Also by command line:

set ray_opaque_background, on
set ray_trace_mode, 1
ray
png picture.png

or:

set ray_trace_mode, 1
png /modesto/strbio/picture.png, width=2000, height=1200, dpi=600, ray=1

Note that objects in a deeper position look lighter. This is the “fog” effect. See: https://pymolwiki.org/index.php/Fog and https://pymolwiki.org/index.php/Depth_cue

Note

If Ray takes too long, you can try ray=0 or use draw: https://pymolwiki.org/index.php/Draw

See more details in the following links:

https://pymolwiki.org/index.php/Ray

https://pymolwiki.org/index.php/Png

2.10 10. BONUS TRACK: More info and useful resources about Pymol.

3 PART B: Paper-style protein figure challenge

Make a ready-to-publish picture of your favorite protein.

As a suggestion, you can reproduce the top panels in the Figure 1B of Gao et al. (2020) or Figure 4b from Cilleros-Rodríguez et al. (2022), but any structure involving more than one domain and/or with a substrate/cofactor molecule can be a good challenge. You may use Pymol or any other application, including Py3Dmol in a Jupyter Notebook.