CHEM 313 Protein Exercise Resource____________________________________________________________________________
This exercise is designed to help you understand and investigate some of the basic physical properties of proteins as determined by their primary sequence. The goals of this exercise are
To use real proteins to explore primary, secondary, tertiary and quaternary structure.
To use bioinformatics databases to find protein information.
To analyze protein primary structure.
To view and observe protein secondary, tertiary and quaternary structure.
To understand that protein primary sequence determines and affects larger protein structures (sequence, shape and function).
To appreciate the diversity of protein structure, function and shape.
You will look up the protein data bank identifier (PDBID) of a particular protein and examine its entry in two databases: UniProt and the Protein Data Bank (PDB). There is a lot of information in these entries that may be unfamiliar to you. Don't panic! You are new to biochemistry and these documents contain a lot of information that is designed for experts. You will be looking for some basic information and this page will help you find it. One of the challenges of this assignment is to examine and explore to find the information you need to complete the assignment. To give you an idea of what you will be seeing and doing see the information below.
For a copy of the assignment handout click here
Looking up your PDBID in UniProt
1. Go to the UniProt Database webite http://www.pir.uniprot.org/search/textSearch.shtml
2. Search the database for a particular UniProtID. To do this pull down the menu under 'Select field and insert query below:" and select :UniProt UIDs/ACs. Your screen should look like this:
3. Enter your accession number into the box and click on "Search"
3. An entry for your protein should come up on the next screen. An example is shown below. Click on the link in the first column to get the entry page for your protein.
4. Use the information in the entry to answer the questions listed in your handout. This document is large and somewhat complicated. Scroll down and look for information you can understand and might need. Take your time and examine the page. For the question that asks you to describe the function of the protein, I would like you to do more than just list or restate what is listed in the 'Function' section. At least try to put it in your own words or get more information so that you can describe what the protein does. If you need to use information from the PDB website (see below) or go someplace else to read a bit on the internet, please do that.
Using the PDB to analyze structure
You can also look up your PDBID in the Protein Data Bank. Here is a link to this website http://www.rcsb.org
Type your PDBID into the search box and press "Site Search"
You should go straight to a summary page for your protein. It looks like this except it has the information for the protein you looked up.
Places to go for useful information
1. The Structure Summary Page It has some information about the name of the protein and the number of subunits. It also has links to other pages with more specific information.
2. The Sequence Details Page (click the "Sequence Details" tab shown below that's near the top of your protein's page)
The page shows a diagram of the primary sequence for the protein with a legend that tells you the type of secondary structure each amino acid is involved in. There are sometimes several things displayed above the structure (domain assignments other notes). You can remove them by looking under the "Image" heading and choosing to "hide" all things but the "Stride Secondary Structure." This makes the map look clearer and it is easier to interpret.
Note: n The STRIDE and DSSP secondary structure views are two different computer algorithms that are used to estimate whether that particular part of the protein is in an alpha helix or beta sheet. The STRIDE algorithm is newer and more accurate so we're using that.
Here's a key for the symbols:
Red squiggles = alpha helix
Blue arrows = beta sheet
Yellow curved lines = random coil
When you are finished with this and want to move along click on the "Structure Summary" tab to return to that page.
3. Viewing Your Protein in 3D: On teh Structure Summary page click the MBT PROTEIN WORKSHOP link under the picture of your protein in the box on the right of the page. This will load a Java applet that will allow you to view your protein's structure in 3D! Be patient. This may take some time to load. When it does the screen looks like this:
How to use this tool:
1. To Move the molecule around: use your mouse to rotate the molecule around and get a better view of the entire molecule.
2. To show protein chains (if it has quaternary structure), ribbons or the atoms and bonds. To do this you:
- Choose #1 "Visibility" as your tool
- Choose #2 "Atoms and Bonds" to show the atoms in the selected chain or "Ribbons" to show a ribbon for that chain.
- In #4 "Choose items" box click on the chain you'd like to effect. You may show the atoms and the ribbon, just the ribbon, just the atoms or you can make a chain disappear if you deselect both the atoms and ribbons.
3. To color the backbone to show information about the protein's primary sequence, secondary structure or the properties of the amino acids use the Shortcuts tab. You will need to do this to show where the hydrophobic amino acids are located for the worksheet. See the photo below to see what this looks like. The more hydrophillic amino acids are shown in blue and those that are more hydrophobic are redder.