Exercise 2.
Structural alignments
Working with 2 or more proteins

1. Load structures
   Load structure 1A6G of sperm whale carbonmonoxy myoglobin (MbCO)
   Load structure 1GDI of lupin carbonmonoxy leghemoglobin (LbCO)
   If you cannot see both proteins click on the "Center" button of the Toolbar
   In Control Panel, click on Header to switch between the 2 loaded structures
   The structure that shows in Header is the structure on top in the Main Window and in Control Panel
   The following 4 steps are to show you the options of the Header buttons
   In Control Panel Header, un-check "Visible" to see the effect of this option
   Place back the checkmark on "Visible"
   Then, click on "Move" and separate the proteins with the 2nd button of Tool Bar
   Place back the checkmark on "Move"
   In Control Panel Windows open the Layers Infos window
   This window provides some reduntant information - typically I keep it closed
2. Alignment Window
   Make visible the Alignment Window
   The top structure is the reference structure (MbCO)
   Place the cursor on top of the first residue of the bottom structure, watch the residue blinking in the Main Window and selected (in red) in Control Panel
3. Fit the coordinates to accomplish superposition of structures
   In Toolbar Fit Magic Fit and select "C-alpha atoms"
   Read the Root Mean Square Deviation (RMSD) in the Tool Bar (you can also calculate it using Fit Calculate RMS
   Click the "Center" button
   Place the cursor on top of residues in the bottom row (2nd structure) of the Alignment Window and check the feedback at the bottom left corner
4. Generate structural alignment
   In Fit click on Generate structural alignment
   Check now the Alignment Window - Structural alignment has been generated with introduction of gaps, etc
   Place the cursor on residues of the bottom structure of the Alignment Window, check the blinking residues, check the RMSD's (they are smaller now)
   Click the little arrow in the Alignment Window to see the threading energy
   In File Save "Alignment" - Save alignment in a text file as junk.txt
   Open junk.txt with a text editor
5. Find a Structural template for a raw sequence
   Launch Deep View
   In Tool Bar Swiss Model Load Raw Sequence to Model use provided file myfile.txt (in Fasta format)
   A model of a linear polypeptide with your sequence will show up in Main Window
   Make sure the following settings are correct in Preferences Swiss Model and Network :
   • Template Server: http://swissmodel.expasy.org/cgi-bin/blastexpdb.cgi
   • Modelling Server: http://swissmodel.expasy.org/cgi-bin/sm-submit-request.cgi
   • Network Server: www.swissmodel.unibas.ch Port: 27000
   • Also your e-mail address and name
   
   Submitt job using Toolbar Swiss Model Find appropriate PDB templates
   Follow instructions
   This is actually the sequence of soybean leghemoglobin and there are several structures available
6. Find a structural template for a mannually optimized sequence
   Launch Deep View
   Read PDB file 1A6G
   Read raw sequence myfile.txt
   Align it mannualy - you can place the cursor in the Alignment Window and introduce or delete gaps using the spacebar or backspace keys
   Submit modeling request in Toolbar Swiss Model Submit modelling request
   Follow instructions
   Wait for 4 e-mail messages among them one with the coordinates in PDB-format file, and one with structure validation
7. Automated structural alignment procedure at the internet
   Go to Swiss Model Server and submit your raw sequence
   You will receive the information via e-mail