User:Rasmusville/sandbox

=PyMOL guide - AlphaDoggies= This guide will give short introduction on how to use PyMOL for creating beautiful figures of proteins. In total we will present three ways in which you can visualize your protein: a general representation of your protein, a representation of the active site of the protein and how you can put together two dimers or subunits and visualize a tetramer or dimer. In all three guides the protein Yeast Alcohol Dehydrogenase I (ADH1) will be used as template

ADH1 is found in Saccharomyces Cerevisiae, which reduces acetaldehyde to ethanol in the fermentation of glucose. The enzyme is present in many organisms and therefore the enzyme has homologs in multiple species. ADH1 consist of four identical subunits each with 347 amino acid residues, the subunits are connected by disulfide bonds at the Cys-277 residue. The 3R/NAD coenzyme (Nicotinamide-8Iodo-Adenine-Dinucleotide) binds to two of the subunits, Trifluoroethanol binds to all four subunits, and zinc ions bind to the oxygen of Trifluoroethanol, changing the conformation before optimal catalytic effect can be observed.

General representation of a protein


This guide will show you how to make a general visual representation of a protein. The end result should be something like figure 1.A in the article. Fetch the protein by typing the following command in the command line of PyMOL: Both dimers will now be shown. To remove one of the dimers use the following commands: Now in the viewer window click on “A” (action) in the bar named “dimer” and then click “remove water” to remove the water molecules. Then click “S” (show) and choose “as” and click “cartoon” to show the dimer as a cartoon instead of a stick model. Then click “C” (color) and choose “by chain” and click “chainbow”. The dimer is now shown in a range of different colors.
 * fetch 4W6Z
 * create dimer=(chain A,B)
 * delete 4W6Z

To show the NAD and TFE and zinc click “S” in the bottom right corner of the viewer window. Now the sequence will be shown over the protein in the viewer window. In the sequence select “8ID” and “ETF”. In the “sele” bar click “S” and choose “as” and click “sticks”. Then click “C” and choose “by element” and click on the fifth color option or whatever you see fit.

To show the zinc molecules select “ZN” in the sequence. In the “sele” bar click “S” and choose “as” and click “spheres”. Then click “C” and click “tv_blue”.

The visual representation of the protein is done. The last thing to do is to adjust the viewing angle in a way you like, set the rendering settings and save the image. A suggestion would be to set antialias to 2 and set the background to be transparent: Now you are able to render your image and save it: You can adjust the minimum number of pixels on one of the dimensions of the picture by typing a number after ‘ray’ e.g. typing ‘ray 1000’ makes the picture dimensions 1000 x above 1000.
 * set antialias, 2
 * set ray_opaque_background, off
 * ray 1000
 * png figure.png

Representation of the acitive site of a protein


This guide will teach you how to find the active site and label the surrounding amino acids. In the article figure 3. The active site of subunit A is depicted. To remake a similar figure, you might be able to find information about the active site on the Uniprot webpage, however, in this case it will be easier to use the information given in the article.

Start by fetching the protein using the PyMOL command line: you get the whole tetramic structure, make sure you can see the sequence by choosing “Display” then “Sequence” or as stated above for figure 1A. Hide one of the dimers.
 * fetch 4W6Z

In the article we see that the 45th and 48th residues (threonine and histidine, respectively) are located in the A-subunit active site.

Choose e.g. His-48 by using “select resi 48” and rename either by clicking the “A” then rename selection or entering “set_name sele,His-48” in the command window. Do the same for Thr-45. Using the A button on either choose “zoom” and you’ll be directed to the site of both residues.

Select 4W6Z and press “Hide – Everything”, then choose His-48 and Thr-45 and press “Show – As – Sticks” for both. Remember, when you choose Show-as, all other setting will disappear. If you press e.g. “Show-Sticks” it will add upon the previous setting. If NAD, Zink and TEF can be found in the end of the 4W6Z sequence as stated for figure 1A. You can create a selection from the sequence or choose to download the SDF files from the PDB webpage of 4W6Z. The ligands can be opened into your current project. NB! Save all your work before making major changes, as not all things can easily be redone.

Ball and stick representation can be made choosing “A – Preset – Ball and Stick” and colors can be changed using “C” e.g. “Color by element”.

Dashed connection lines can be made e.g. by selecting the molecules you want to find connections between and then “A – Find – Polar contacts – Within Selection”.

The amino acid residues can be labeled by using the “L” (Label) button and press residues. To move the labels click on the “Mouse Mode” text in the lower right corner and make sure it says “3-button editing”. Then if holding down ctrl+left-click or ctrl-shift-left click the label can be moved in x, y and z axes. Otherwise command the following command can be used: where x, y and z are the distances in the x-, y- and z-axis you wish to move the label.
 * set label_position,(x,y,z)

Save a picture by the using the directions stated earlier.

Representation of a tetramer or a dimer


This guide will teach you how to rotate and move two dimers (or two subunits) to form a tetramer (or a dimer) along with adjusting visual settings and rendering. The end goal is to have a image resembling figure 1.B in the article. The first step is to fetch the protein structure into PyMOL by typing the following command in the command line: Next you need to split up the protein into its two dimers: The protein itself can be deleted as it will not be used anymore. The two dimers are not put together in a tetramer which needs to be done manually. We orient the dimers and set the center of rotation to dimer1 (the dimer you wish to adjust): dimer1 is now ready to be rotated and moved. First of all it is rotated 180 degrees around the y-axis and afterwards it is rotated 70 degrees around the x-axis: dimer1 is now rotated correctly. The next step is to move dimer1 closer to dimer2. dimer1 is moved a distance of -35 in the X-axis: In order to get a correct alignment for your dimer or tetramer you might need to fiddle around with the rotation and movement of your own subunits or dimers.
 * fetch 4W6Z
 * create dimer1=(chain A,B)
 * create dimer2=(chain C,D)
 * delete 4W6Z
 * orient
 * origin dimer1
 * rotate y, 180, dimer1
 * rotate x, 70, dimer1
 * translate [-35, 0, 0], dimer1

The tetramer is now correctly aligned and the visual settings needs to be adjusted in order to get a nice picture of the tetramer. The line representation is hidden and the cartoon representation is shown. Additionally, for dimer1 the cartoon representation is set to be of the type ‘loop’: The following visual settings were also adjusted: The visual representation of the protein is now set. However, water molecules are not removed and the visual representation of zink molecules as well as 8ID and ETF is not optimal. The water molecules are selected and hidden: Then the zink molecules is selected and set to be shown as spheres: and 8ID and ETF are selected together and set to be shown as sticks: Last but not least all colors of the figure are set: The visual representation of the tetramer is done and an image of the tetramer can be saved as stated in the end of the section General representation of protein.
 * hide lines, all
 * show cartoon, all
 * cartoon loop, dimer1
 * set cartoon_smooth_loops, on
 * set cartoon_oval_length, 1
 * set cartoon_rect_length, 1
 * set cartoon_transparency, 0.5, dimer1
 * select water, resn HOH
 * hide (water)
 * select zink, resn ZN
 * show spheres, (zink)
 * select substrates, resn 8ID or resn ETF
 * show sticks, (substrates)
 * color lightblue, (chain A)
 * color lightmagenta, (chain B)
 * color blue, (chain C)
 * color magenta, (chain D)
 * color grey70, (zink)
 * color green, (substrates)