1. SMART Teams: Students Modeling A Research Topic Jmol Training 101!

2. There are three methods that may be used to determine structures deposited in the protein data bank: X-ray Crystallography Structure 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

3. 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

4. X-Ray Crystallography

5. Crystal of the Molecule of Interest

6. X-Ray Crystallography X-Ray Crystal of the Molecule of Interest

7. X-Ray Crystallography X-Ray Crystal of the Molecule of Interest X-ray film

8. X-Ray Crystallography Series of Computations X-Ray Crystal of the Molecule of Interest

9. X-Ray Crystallography Defines the X, Y, and Z coordinates for each atom in a molecule Series of Computations X-Ray Molecule of Interest

10. 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 What do you see?

11. Building a Model What you see + What you think = What you get

12. Final Model 3D models take visual representation to a new level of awesomeness!

13. 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.

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

15. Cryo-EM

16. 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 File Copy and paste the pdb file of the protein to be modeled (Instructions to follow)

17. 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 Retrieve a PDB File

18. The Protein Databank Molecule of the Month! Search Bar

19. PDB File Search You may search by: Search Bar Molecule name Scientist name File name

20. 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

21. Primary Citation You 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

22. More Summary Tab Information Method Scroll down the screen while on the “Summary” tab to find information about: Ligands Structure Determination Method Ligand

23. 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

24. 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

25. Opening Your File in Jmol Open Jmol.jar

26. Opening Your File in Jmol Your screen should look like this: Model Design Box Console Box: Type your commands here

27. Opening Your File in Jmol Drag and drop the PDB File into the black screen.

28. You Are Ready to Design! The Jmol Quick Reference Sheet is your lifeline for protein model design!

29. You Are Ready to Design! Try the Mouse Movements!

30. 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)

31. 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

32. 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

33. 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

34. Highlighting a Specific Chain To select a specific section of a molecule, type: select :b and backbone To color the backbone, type: color green

35. Highlighting a Specific Chain To select a specific section of a molecule, type: select :a and backbone To color the backbone, type: color yellow

36. Highlighting Secondary Structures To highlight secondary structures, type: select :a and backbone and helix color red select :a and backbone and sheets color blue

37. Adding a Sidechain To Display Sidechains Refer 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 1.25 wireframe 1.0

38. Adding a Sidechain Click on the amino acid and check the information provided in the script console box.

39. 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!

40. 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

41. Coloring a Sidechain To color only the sidechain and not the backbone, type: select glu177 and sidechain color cpk

42. 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

43. 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 sheets calculate hbonds hbonds 1.0 set hbonds solid set hbonds backbone color hbonds

44. Adding Disulfide Bonds Disulfide bonds can be an important part of the molecular story of your protein. To display disulfide bonds, type: select all ssbonds on ssbonds 1.0 set ssbonds backbone color ssbonds purple You may also chose to highlight the cysteine sidechains responsible for forming the disulfide bond instead.

45. 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 all calculate struts struts 1.0 color struts tan

46. 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

47. Opening Your Saved Work To Open: Your folder MUST have PDB file Shortcut to Jmol *.jpg