These are exercises and for the most part not questions to be answered. The ones that you should answer are in pink:

1. Getting to know SPDBV

Save 1HEW.pdb onto your computer - Open in chime (from Netscape) and save pdb file from the chime files menu or by long-clicking here – if you choose the later option, you need to open the file in word and save it under a different name in text format)

Start SPdbV

load 1HEW.pdb (hit return)

click the right mouse button (= sign on Mac, in most other instances the alt-key together with the mouse-click has the same effect as the right button on the PC mouse) to center the molecule

click on the three cursor control buttoms and rotate/move/enlarge lysozyme picture

click on the page icon and go through the pdb file

open the control window (Window -menu).

open the align window (Window -menu)

select all (Select menu)

Window Ramachandran plot (In case you forgot what this is about, go here.)

in the control or the align window select different residues

select all (Select menu)

Explore different coloring schemes (CPK, secondary structure, accessibility) and display options (show CA trace only, show oxygen, …)

REMARK: If you do serious work save your work periodically, sometimes it is impossible to recover from an inadvertent mouse click)

Select the NAG inhibitor (go to the at the bottom of the control panel, click with left mouse button).

Color CPK

Invert selection (return) (Select menu)

Color secondary structure

Invert selection (return)

Tools calculate H-Bonds

right click inside the side chain column in the DISPLAY window to turn off side chain display

select Neighbors of selected aa (5 to 10 Angstrom is appropriate) (Select menu)

hit return

click on side-heading in control panel (clicking in the header acts only on selected residues), add labels, change color ….

select group properties non-polar aa (Select menu)

click on Header COL in control panel select blue color to color hydrophobic residues blue

Are there “blue” residues interacting with the N-Acetyl glucosamines? How come? (Aren’t carbohydrates supposed to be hydrophilic?)

The resulting display after some manipulations might look like this (after you save it as a pov-ray image):

yellow: the NAG inhibitor;
blue: residues in the binding pocket that are non-polar, depicted as space filling balls;
red: other amino acids in the binding pocket;
gray: the rest of the Lysozyme molecule, but only the backbone.

Play around, if in doubt use the ? buttom. 

The worst that can happen is that you'll have to restart your computer.

Open the alignment window and display the complete lysozyme molecule. Observe the color change in the structure that happens when you move the mouse over the sequence in the alignment window.

Other things to try:
  3D rendition (in the display menu),
  slab view (shift and mouse move the slab),
  explore the make up of the pdp file (text icon below the cursor control),
  have a look at the opening control window (upper left icon below the cursor control).

If you right click on a residue in either the alignment window or the control window, the display centers on this residue.

Control and mouse click adds residues to the list of selected residues (works in either window)

2. Exercise on multi subunit proteins:
Aligning F-ATPase alpha and beta subunits

Start SPdbV

Open 1bmf.pdb (save from here - remember the text file re-saving from above -, or open with chime – netscape - and save from file menu)

(F-ATPases are typically found in bacteria and mitochondria and plastids. They either function as proton pumping ATPases, or they use the proton gradient to synthesize ATP. They can be dissociated into two parts; the F1 portion binds and hydrolyses ATP. The conformation change associated with this catalytic cycle of ATP binding, hydrolyses and ADP+Pi dissociation is communicated to the membrane imbedded F_O-portion via the gamma subunit (part of F1). Click here for animated gifs displaying the catalytic cycle, and here for a movie of the actual rotation as seen under the microscope)

Color Chain

Change color chainD to grey/blue (to do this right click in control panel on D in first column to select chain D, right click on COL, select color)

Scroll down the control panel and select all ATP analogs (press ctrl key and right click to select)

right click on COL in heading and select red color

Read the pdb file to get info on which chain is which

select chain F (including nuc) and save selected residues as betaTP.

select chain A (including nuc) and save selected residues as alphaE.

After playing with the F1-ATPase, close this file and open betaTP and alphaE.

Display layer info (windows menu)

To avoid confusion select and display the nucleotides only

There are different ways to align 3-D structures. One way is to select 3 corresponding points in each of the two structures. To do so you can use the substrate molecule.

Using the mov check off in the Layer Info, reorient the two ANPs so that they are in a similar orientation (but not overlapping).

Click on the align bottom with the 3 green and 3 red dots. Notice the red instructions that appear in the header next to the pdb-page icon. Follow these instruction using three corresponding atoms.

SHIFT DISPLAY Show CA trace only (Shift makes the commands act on both layers)

Using the mov checks in the Layer info, move the two chains next to each other.

What do you think about the result?

Another way to align structures is to use the magic fit in the tools command. Do this and run improve fit (notice the red info in the header)

Click on alpha in Layer info to make the alpha subunit the active layer

Color CPK

Make the beta subunit the active layer

COLOR rms . The further the atoms in the beta subunit are away from the alpha subunit, the longer wavelengths it is the colored.

DISPLAY Show alignment window - gives you the aligned sequences.

Use the mouse over function to find the parts of the structure which are used in the alignment, and the part that is not used. What could one do to get an alignment of the carboxy portion parts that is not aligned after magic fit?

How does this alignment compare to the ones calculated using Clustalw? (You can save your alignment as a text file from the File menu; testseq2 in Assignments4 contained both beta and alpha F-ATPase subunits.)

If you have time, repeat the exercise for the four histones that are part of the nucleosome (see lecture notes on class 5).