SMART Teams:Students Modeling A Research Topic

Jmol Training 101!

There are three methods that may be used to determine structures deposited in the protein data bank:

X-ray CrystallographyStructure must be in a crystallized form

Nuclear Magnetic Resonance (NMR)Structure may be in solution

Cryo-electron microscopy (Cryo-EM)Used primarily for large multiprotein assemblies

Methods Used to Determine Molecular Structures

X-Ray Crystallography

Determination of the three-dimensional structure of molecules by means of diffraction patterns produced by x-rays of crystals of the molecules rr

X-Ray Crystallography

X-Ray Crystallography

Crystal of the Molecule of Interest

X-Ray Crystallography

X-Ray

Crystal of the Molecule of Interest

X-Ray Crystallography

X-Ray

Crystal of the Molecule of Interest

X-ray film

X-Ray Crystallography

Series of Computations

X-Ray

Crystal of the Molecule of Interest

X-Ray Crystallography

Defines the X, Y, and Z coordinates for each atom in a molecule

Series of Computations

X-Ray

Molecule of Interest

What Do You See?

• collect images around crystal to record the position of all the spots

• spots represent where x-rays were deflected by electrons in the molecule

• resolution is reflection of how well ordered the atoms are in the crystal

phenylalanine

Building a Model

What you see + What you think = What you get

Final Model

3D models take visual representation toa new level of awesomeness!

The Case of the Missing Hand

• Sometimes a clear model is difficult to produce.• Uncertainty may occur in

interpreting data collected from a crystal if a portion of the molecule has multiple conformations.

For example, the photograph on the left demonstrates the blurry image that is produced if a student moves their hand while being photographed.

NMR

http://phet.colorado.edu/en/simulation/mri

Cryo-EM

Let’s Begin Jmol: Create a Folder

First create a folder to store the information needed to design your model.

Include a Copy of Jmol Copy and paste a copy of the Jmol.jar file into the folder you created

Include the PDB FileCopy and paste the pdb file of the protein to be modeled (Instructions to follow)

Retrieve a PDB File

The Protein Data Bank (pdb) is the worldwide repository for the processing and distribution of 3-D biological macromolecular structure data.

http://www.pdb.org

The Protein Databank

Molecule of the Month!

Search Bar

PDB File Search

You may search by:

Search Bar

• Molecule name• Scientist name• File name

PDB File Download

File Name

• Type the file name in the search bar. We will use 2AAI.• Click on the Download Files and select PDB File (Text)• Place this file in the folder that you have created

Primary CitationYou may find the following information on the “Summary” tab:

• Title of the primary citation• Authors of the primary citation• Journal in which primary citation was published• Abstract

More Summary Tab Information

Method

Scroll down the screen while on the “Summary” tab to find information about:• Ligands• Structure Determination Method

Ligand

PDB File Search

Click on the “Sequence” tab.Information about primary and secondary structure of the protein may be found here!

Primary and Secondary Structures

Jmol Version

Be sure to use the correct version of Jmol!

We will not be able to view your files if you use a version other that the one specified!

This year we will use version:

Jmol 14.2.15

Opening Your File in Jmol

•Open Jmol.jar

Opening Your File in Jmol

•Your screen should look like this:

Model Design BoxConsole Box:

Type your commands here

Opening Your File in Jmol

Drag and drop the PDB File into the black screen.

You Are Ready to Design!

The Jmol Quick Reference Sheet is your lifeline for protein model design!

You Are Ready to Design!

Try the Mouse Movements!

Changing the Background Color

Type all commands into the console box.

To change the background color, type:background white

(you may want to try a few other colors)

Jmol Colors

Jmol Colors

To explore other Jmol color options you can investigate the following site: http://jmol.sourceforge.net/jscolors

Scroll down until you see: JavaScript Colors

Alpha-Carbon Backbone Format

Type commands in the console box.

To display the alpha carbon backbone, type:backbone 1.5

You will notice a “funny line” in the backbone – the cartoon display is still turned on. Type:cartoon off

Restrict Command

To “get rid of the water molecules”, type:restrict not water

Notice the other ligands are still displayed. To remove them, type:restrict not hetero

Highlighting a Specific Chain

To select a specific section of a molecule, type: select :b and backbone

To color the backbone, type: color green

Highlighting a Specific Chain

To select a specific section of a molecule, type: select :a and backbone

To color the backbone, type:color yellow

Highlighting Secondary Structures

To highlight secondary structures, type:select :a and backbone and helixcolor red

select :a and backbone and sheetscolor blue

Adding a Sidechain

To Display SidechainsRefer to the primary citation to determine the relevant sidechains to display in your model design.

One such side chain in ricin is glutamic acid 177, as it is a key catalytic amino acid in the active site.

To display this sidechain, type:select glu177 and (sidechain or alpha)spacefill 1.25wireframe 1.0

Adding a Sidechain

Click on the amino acid and check the information provided in the script console box.

Adding a Sidechain

If you do not include (sidechain or alpha) you will get what is known as a “bumpy backbone”, shown to the right.

AVOID the dreaded bumpy backbone at all costs!

Adding a Sidechain

AVOID the dreaded bumpy backbone and add Arginine 180 to your practice model!

$ select arg180 and (sidechain or alpha)8 atoms selected$ wireframe 1.0$ spacefill 1.25

[ARG]180:A.NH2 #1417 20.909 64.017 53.14

Coloring a Sidechain

To color only the sidechain and not the backbone, type:select glu177 and sidechaincolor cpk

Coloring a Sidechain

A common error is to color the backbone as well as the sidechain (shown at right).

Avoid this error by using the commands on the previous slide.

Now correctly color arg180

Adding Hydrogen Bonds

In some models you will want to display the hydrogen bonds (especially in the beta sheets)

To add hydrogen bonds to beta sheets, type:select sheetscalculate hbondshbonds 1.0set hbonds solidset hbonds backbonecolor hbonds <lemonchiffon>

Adding Disulfide Bonds

Disulfide bonds can be an important part of the molecular story of your protein.

•To display disulfide bonds, type:select allssbonds onssbonds 1.0set ssbonds backbonecolor ssbonds purple

You may also chose to highlight the cysteine sidechains responsible for forming the disulfide bond instead.

Adding Struts

In some models you will need to stabilize the model or connect a ligand to the model. To do this you will need to add struts.

To add struts, type:select allcalculate strutsstruts 1.0color struts tan

Saving Your Work

Be sure to save your work!

To save:

Click on the camera icon

Name your file

It should now be saved in the folder you created

Opening Your Saved Work

To Open:

Your folder MUST have

• PDB file

• Shortcut to Jmol

• *.jpg