HONORS OPTION:  GEN CHEM 1

Fall 2006

(self note:  contact Jason Cross to ensure usrname/passwd correct and to ensure access to /common folder.)

Part 1:  Proteins and Enzymes - on PC Network

• Some small molecules:  Water |  Ethanol |  A Collection

• A typical protein: Triose Phosphate Isomerase

• A complex of two proteins:  Thrombin and Hirudin

Part 2:  Intermolecular Forces - IMFs

Part 3:  HIV Protease: an Enzyme at Work - Video

Part 4: Molecular Modeling of the
HIV Protease/Inhibitor Complex

In this exercise, you will study the structures of HIV protease and an inhibitor which binds to the protease. You will use a powerful molecular modeling program called VMD, which was developed at the University Illinois (Champaign/Urbana).  For more information, see the website below

• VMD Homepage, University Illinois

OpeningVMD

First you must log into the Silicon Graphics network.  For a user name, type in final01 up to final12  as assigned in lab.  The password is: f03.final.  On the top menu bar, select Applications, Chemistry, VMD.  Three windows will open for you. 

1. VMD Main - contains menu bar for running VMD
2. VMD 1.8.1 OpenGL Display - displays molecules
3. VMD - contains text and place for command line input

Loading a Molecule

Our first step is to load our molecule. A pdb file, 1HSG.pdb, that contains the atom coordinates of HIV protease and an inhibitor is provided with the tutorial.
 

1

Choose the File New Molecule... menu item Fig 1(a) in the VMD Main window. Another window, the Molecule File Browser (b), will appear in your screen.

2

Click Up One Directory until the list of folders doesn't change anymore. The scroll down the list and select with your left mouse button /usr.  A new list of folders will appear.  Select /people and continue with the following selections:  /faculty then /chem  then /hjakubow .  At this point no files are shown but rather the work Permission Denied.    In the line that shows the path, type in immediately after hjakubow (with no spaces) /common.  A list of files should now appear.  Select 1HSG.pdb and then Open.   

Note that when you select the file, you will be back in the Molecule File Browser window. In order to actually load the file you have to press Load (d). Do not forget to do this! 

Now, HIV protease is shown in your screen in the OpenGL Display window. You may close the Molecule File Browser window at any time.   You will see many red dots around the protein.  These represent the oxygen of water molecules that co-crystallized with the protein that was used to determine the x-ray structure.
 

Displaying the Protein

In order to see the 3D structure of our protein we will use the mouse and its multiple modes.

1

While holding the left button pressed over the protein in the OpenGL Display, move the mouse and explore what happens. This is the rotation mode of the mouse and allows you to rotate the molecule around an axis parallel to the screen Fig. 2(a).

2

If you press the second button and repeat the previous step, the rotation will be done around an axis perpendicular to your screen (b) (For Mac users, the second button is equivalent to press the command key while holding the mouse button pressed).

3

In the VMD Main window, look at the Mouse menu (Fig 3). Here, you will be able to switch the mouse mode from Rotation to Translation or Scale modes.

4

The Translation mode will allow you to move the molecule around the screen while holding the first button pressed.

5

The Scale mode will allow you to zoom in or out by moving the mouse horizontally while holding the first button pressed.

It should be noted that the previous actions performed with the mouse do not change the actual coordinates of the molecule atoms.

Another useful option is the Mouse Center menu item. It allows you to specify the point around which rotations are done.
 

6

Select the Center menu item and pick one atom at one of the ends of the protein. (The cursor should display a cross.)

7

Now, press r, rotate the molecule with the mouse and see how your molecule moves around the point you have selected.

Exploring Different Drawing Styles

VMD can display your molecule using a wide variety of drawing styles. Here, we will explore those that can help you to identify different structures in the protein.
 

1

Choose the Graphics Representations... menu item. A window called Graphical Representations will appear and you will see in yellow Fig 4(a) the current graphical representation used to display your molecule.

2

In the Draw Style tab (b) we can change the style (d) and color (c) of the representation. In this section we will focus in the drawing style (the default is Lines).

3

Each drawing style has its own parameters. For instance, change the Thickness of the lines by using the controls on the right bottom part (e) of the Graphical Representation window.

4

Now, choose from Drawing Method the VDW (van der Waals) menu item. Each atom is now represented by a sphere. In this way you can see more easily the volumetric distribution of the protein.

5

In order to see the arrangements of atoms in the interior of the protein, use the new controls on the right bottom part of the window (e) to change the Sphere Scale to 0.5 and the Sphere Resolution to 13. Be aware that the higher the resolution you choose, the slower the display of your molecule will be.

6

Note that in the Name coloring method, each atom has its own color, i.e: O is red, N is blue, C is cyan and S is yellow.

7

Press the Default button. This allows you to return to the default properties of the drawing method.

The previous representations allows you to see the micromolecular details of your protein. However, more general structural properties can be seen by using more abstract drawing methods.
 

The last drawing method we will explore here is called Cartoon. It gives a simplified representation of a protein based in its secondary structure. Helices are drawn as cylinders, β sheets as solid ribbons and all other structures as a tube. This is probably the most popular drawing method to view the overall architecture of a protein.
 

10

Choose the Cartoon style and set the Beta Sheet Thickness as 3, the Helix/Coil Radius as 1.5.

11

Identify now how many helices, betasheets and coils are present in the protein.

Exploring Different Coloring Methods

1

Now, let's modify the colors of our representation. Choose the ResType coloring method Fig. 4(c). This allows you to distinguish non-polar residues (white), basic residues (blue), acidic residues (red) and polar residues (green).

2

Select now the Structure coloring method (c) and confirm that the cartoon representation displays colors consistent with secondary structure.

Exploring Different Selections

Let's look at different independent (and interesting) parts of our molecule.
 

1

In the Selected Atoms text entry Fig. 4(f) of the Graphical Representations window delete the word all, type helix and press the Apply button or hit the Enter key. (Do this every time you type something.) VMD will show just the helices present in our molecule.

2

In the Graphical Representations window choose the Selections tab Fig. 6(a). In section Singlewords (b) you will find a list of possible selections you can type. For instance, try to display β sheets instead of helices by typing the appropriate word in the Selected Atoms text entry.

Combinations of boolean operators can also be used when writing a selection.
 

3

In order to see all that is not helix and not β sheet, type the following (not helix)and(not betasheet)

4

In the section Keyword (c) of the Selections tab (a) you can see properties that can be used to select parts of a protein with their possible values. Look at possible values of the Keyword resname (d). Display all the Lysines and Glycines presents in the protein by typing (resname LYS)or(resname GLY). 

5

Now, change the current representation to CPK style and the coloring method to ResID by using the previous described buttons in the Drawing Method (Style) tab. In the screen you will be able to see the different Lysines and Glycines. How many of each one can you see?

The button Create Rep Fig 7(a) in the Graphical Representations window allows you to create multiple representations and therefore have a mixture of different selections with different styles and colors, all displayed at the same time.
 

1

Be sure that the current representation is in CPK style and coloring method Name

2

Set the current selection as protein.

3

Press the Create Rep button. Now, using the menu items of the Draw Style tab and the Selected Atoms text entry, modify the new representation in order to get Ribbons as the drawing method, Structure as the coloring method, and helix as the current selection.   Call this newly created representation b:

Figure 7: Multiple Representations of Ubiquitin (not the protein we are studying).

4

Repeating the previous procedure, create the following three new representations:

The complex you have seen above, no matter how you render it, is still quite complicated.  We have used a program called MOE (available on the SGI network) to create a file (HIVMOECONTACTpolarH.pdb) that shows only the inhibitor and the amino acids around it that make contacts.   Load it now.

• select from the top menu bar of the VMD Main windowFile New Molecule
• In the Filename: box, type in the following:

/usr/people/faculty/chem/hjakubow/common/HIVMOECONTACTpolarH.pdb
(make sure you use capitals letters when for HIVMOECONTACT)

What properties must the inhibitor and binding pocket of the enzyme have in common to allow inhibitor binding?  You will soon cover this topic in General Chemistry (Chapter 10, Section 10.2).  In essence there are intermolecular forces - forces between molecules that are distinguished from covalent bonds within a molecule) that allow reversible binding.  In general, there are 3 types of intermolecular forces between the inhibitor and protease (the two different molecules) that may be involved here: