Assignment 1

Your name:
Your email address:

Preparation for future classes

You'll need to set up an account on the UConn BBC** cluster for future classes, so we'll do that first. (If you're interested in what a "cluster" is, see What is a cluster?.) To do this, fill out the following form — Inquiry Selection is "Account for Course" and Course is "Introduction to Molecular Evolution and Bioinformatics (MCB3421)". (It seems that the fields "Department" and "Principal Investigator's Name" may also be required to successfully submit the form — you can get creative... or just use MCB/JohannPeter Gogarten.)


** not that BBC

(This is background information for you to read and explore. You do not need to follow any of the links listed in the introduction, but these might come in handy, if you want to install software on your own computer.)

There are several programs that allow the inspection and manipulation of 3-D structural protein data. In this course we use the swiss protein data bank viewer, aka Deep View. An alternative, very popular to generate rotating or rocking images is pymol . A very simple get to know pymol exercise is here. (If you think protein structures are in your future, you might want to give this a try in your own time).

SPDBV is a good choice, because it also provides an interface to the Swiss Protein databank modeling software.

The SPDBV program is available free of charge from the Swiss Institute of Bioinformatics.

There are several on-line tutorials available to learn the use spdbv:

The exercise in this section is taken with slight modifications from Gale Rhodes' basic tutorial, many of the exercises in the following sections parallel exercises in the basic tutorial.

You can retrieve pdb files from the NCBI, or from the protein structure data bank at Rutgers University. (To do so search for the file -use only the name, omit the extension-, once the page for the structure has loaded, click the download link in the upper right hand corner and click on the link that indicates to download the uncompressed pdb file.) (The ones used in the course are also available here - we will use 1HEW.pdb and 1bmf.pdb today)).


For the instructor to get to know you, and to allow a more organized way to help students, please write your name on a small piece of paper (or on an the wings of an origami crane). If you have a question and the instructor is busy helping other students, please move your name card onto the top of your screen, the instructors will try to help you ASAP.

Exercise 1 :

Do the following:

copy the files labeled 1bmf and 1HEW from here into a folder on the desktop of your computer
(go to the listing, right-click on the name and "Save target as..." to a folder on your computer).

Find "Swiss-PdbViewer" in the Start—All Programs menu to start SPDBV. Close the "About Swiss-PdbViewer" window.

load 1HEW.pdb (click on file, then open PDB file, then navigate to your folder on the Desktop that contains the 1HEW.PDB file). Close the log window that pops up.

The tool bar panel looks a bit like this: (note that this image is from the Mac version -- on Windows a "File Edit Select...Help" menu bar is incorporated into the tool bar panel [on the Mac, "File Edit Select...Help" would appear separately at the top of the screen])

The button which is second from the left allows you to move the structure in the structure window. Click on it and then click and hold in the structure window to drag the protein to the desired location. The third button from the left allows you to zoom in or out. Click on it and then click and hold in the structure window to drag the protein larger or smaller. The fourth button from the left is the rotate button. Click on it and then click and hold on the structure window to rotate the view.

The leftmost button centers the structure in the structure window. Click it now to return the molecule to the center of your screen.

Explore Different Display Options. For large structures, render in 3D may slow down the program, but for small ones like 1HEW, render in solid 3D is a useful choice.

click on the page icon (in the second row of the tool bar and is a white rectangle-- you may need to expand the menu window to see the page icon) and scan through the pdb file.

The Control Panel looks similar to this: (again, this is the Mac version)

the control panel is probably already visible, or you can open the control panel (in the WIND-menu from the tool bar panel).

open the alignment window (in the WIND-menu from the tool bar panel)

select all (look under the select menu).

in the WIND-menu, click Ramachandran plot

in the control panel, select different residues (click on the 'h' and 's' in the first column, then hit return (return make the selected residues visible, else the visibility and the selected residues can be different!). How does the display change in the Ramachandran plot? In the main window? (For more info on the Ramachandran plot see

Your answer --->

select all, hit return

Explore different coloring using the color menu at the very top of the screen (CPK, secondary structure, accessibility) and display options in the display menu at the top of the screen (show CA trace only, show oxygen, ...)
Often the amount of information is overwhelming.

REMARK: If you do serious work save your work periodically, sometimes it is impossible to recover from inadvertent mouse clicks). If you used the interactive feature of the Ramachandran plot to (inadvertently) modify the structure, re-load the original 1HEW file.

Remark2: There is a difference between select (the residue turns red in the control panel) and actually seeing the residue in the main window. If you hit return the selected residues become visible.

Remark3: Especially when working with large structures, it is beneficial to only display the alpha carbons of the backbone. To do so, shift mouse click in the side (side chain) column of the control panel (to turn on/off all checkmarks in this column). In the display menu uncheck "show backbone oxygens", and check "show backbone as alpha carbon trace".

Remark4: Another way to have an overview picture is to select ribbon representation in the control panel. Uncheck (shift mouse click) all other columns and check the rbn (ribbon) column. To have color commands act on the ribbon display, you need to select the ribbon as target for the color commands (either via the first item in the color menu, or via the little black triangle below the "col" (color) header in the control panel.

To highlight residues in the substrate binding pocket do the following:

Select (point the cursor over the NAG201..NAG203 at the bottom of the control panel) the NAG inhibitor (click NAG201, then shift-click NAG203 (select a range)... or click NAG201, then control click adds NAG202,NAG203 to the selection).

Color CPK

Invert [inverse] selection (in the SELECT menu at the top of the screen)

Color secondary structure

In the Tools menu at the top of the screen, choose compute H-Bonds

click "side -" column in the control panel to turn the sidechain display off

Invert selection (only the NAG inhibitor should now show as selected in the control panel)

In the select menu at the top of the screen, select Neighbors of selected residues - check select add to selection button

hit return

click on the + under the "side" header in control panel (acts only on selected residues)

In the select menu at the top of the screen, select group property, and then Non-polar aa

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

Are there "blue" residues interacting with the N-Acetyl glucosamines? How come? (Rotate the molecule to get a good look at things.)

Your answer --->

Can you locate which one of the Tryptophan residues "sits under" the second of the N-Acetyl glucosamines? (tim's hint for the truly desperate: the LABL column in control panel may be of some use, or change the color, or even var der Waals dot spheres [::v] — all these can be turned on/off for individual residues)

Your answer --->

Play around, if in doubt use the ? button. 

The worst that can happen is that you'll have to restart your computer. SPDBV allows you to model protein structures, i.e. you can actually change the structure (e.g., you can drag a dot in the Ramachandran plot; this changes the structure irrevocably). If this happens to you by accident, close the structure and reload. If you work on something important save you work frequently.

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.

The resulting display after some beautifications might look like this:

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.

Trouble shooting: In case your cartoon (ribbon) display does not look nice:
  1st: in the Control panel window, check that the coloring commands are selected to pertain to the the ribbon.
  2nd: under preferences, select ribbon, and place a check mark in the field "render as solid ribbon"


Other things to try:
   3D rendition (in the display menu),
   slab view (shift and mouse forward/backward move the slab through the molecule, shift and mouse left/right change the slab size),
   explore the make up of the PDB file (text icon below the cursor control).

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

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

Can you obtain a figure similar to the one below?

Go to the control panel click on the little black triangle to the right of the col column and select color ribbon, then secondary structure in the color menu. Display ribbon in the control panel, remove the other displays .....



If you have time, do the following exercise, we will return to this next week!

Exercise 2:

Aligning F-ATPase alpha and beta subunits

Start SPdbV

Open 1bmf.pdb

Color Chain

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

Scroll down the control panel and select all ATP analogs (labeled as ANP, ADP or ATP)(press ctrl key and left click to select)

left 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 file - save selected residues as betaTP.

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

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

In WIND(OW) menu - Display layer info

select and display only the nucleotides (ANP600... these may show up at the bottom of each listing as simply "d600")

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 window, reorient the two AMPs so that they are in a similar orientation (but not overlapping).

Click on the align button with the 3 green and 3 red dots (third square from right). Notice the red instructions that appear in the header next to the pdb-page icon. Follow these instruction using three corresponding atoms.

Make all of the aa visible (using the control panel).

SHIFT DISPLAY Show backbone as CarbonAlpha trace chain (Shift makes the commands act on both layers)

Color according to secondary structure. (Alternatively, you can use the ribbon display, colored according to secondary structure.)

Pressing control tab switches the display between the two layers.

(You also could rotate the subunits into a nice position and then 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 fit menu. 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.

WINDOW display alignment window - gives you the aligned sequences.


Which part of the molecule looks different between the Alpha vs. the Beta subunit?

Your answer --->

Is the Walker motif (G--G--GKT) well aligned in the structures? How did you locate the molecule? (tim's hint for the truly desperate: it turns out that there's a really easy way to do this in the menu... however, if you're stuck for ideas, then looking for aa residues near the substrate sounds reasonable, or... about 1/3 of the way scolling along the alignment window, or on line 4 (out of 10) of the alignment text when one clicks on the page icon in the alignment window)

Your answer --->


Finished? Leaving the lab? Head spinning? Remember to...

Choose one of the following options, then click the button.

Send email to your instructor (and yourself) upon submit
Send email to yourself only upon submit (as a backup)
Show summary upon submit but do not send email to anyone.