In this exercise you will learn how to use the Cn3D structure tool to visually display structure files, as well as to do protein structure alignments. Additionally, we'll use RasMol and Swiss Prot Data Bank Viewer (SPDBV) to display files. You'll need to have all three of these applications installed on your computer. For this exercise, we'll assume only RasMol and Cn3D have been installed, but an exercise involving Swiss-Model First Approach will be done to generate a PDB file that you can view using RasMol.
Software:
You'll need to download and install each of these applications, so let's do that first.
Cn3D - Download and install Cn3D 4.1 The file is roughly 2.5 Mbytes and the installation is easy and quite stable. Make sure to read through the tutorial too! (Note that the tutorial explains how to do a pairwise BLAST within the alignment exercise, which is automatic in Cn3D version 4.1)
RasMol - Download and install RasMol. There is a help file that can (and should) be downloaded, and configured when you install RasMol 2.7
SPDBV - Download and install SPDBV. There is a lot to this website, which is actually a 'protein structure portal'. Take your time to familiarize yourself with the the 'First Approach Mode' and then the site for downloading and installing the 'DeepView' tool. Note that this site is in frames.
NCBI Molecular Modeling Database
Next we'll explore the NCBI Molecular Modeling Database (MMDB) as a resource of protein structure related information, including the Conserved Domain Database (CDD).
We'll start our structure journey by searching NCBI's structure database for the gene / protein named 'COMT'
Go to http://www.ncbi.nlm.nih.gov/Structure/ From this link we will search both using the name 'COMT' in the larger rectangular search field, and later we'll come back and search using a PDB id (1VID) in the smaller rectangular search box (Structure summary PDB/MMDB-Id:)
When you type in COMT at the Structure search bar, you'll be directed to a page with structure entries for 1KYZ, 1KYW, 1JR4, and 1VID.
Choose the link to 1VID. When you search for 1VID directly from the Structure page, you go directly to the MMDB Structure Summary.
Both approaches will eventually get you to the same place, the MMDB Structure Summary for 1VID. Let's explore the 1VID MMDB entry!
NCBI Molecular Modeling Database - 1VID
There are four links to be explored here. The two text links on the left, labeled Protein and CDs are first. Follow the Protein link to the 1VID entry in GenPept. Take a minute to read that file entry. The 'accession' number here is the PDB id, and is the primary key for this database entry.
The CDs pfam entry takes you to the Conserved Domain summary, which can be further navigated to the Pfam Conserved Domain Database. Try it! Now go back to the Methyltransf_3 image in the MMDB summary, and click that image. It will take you to the link that we visited above!
Last, go back to the MMDB summary, and then try the chain link , which will show you the structural neighbors to 1VID, which you can import into Cn3D and do VAST alignments. But let's take this a step at a time. Explore some of 1VID's neighbors - why (and how) are they similar to 1VID?
(You can load any of these structures into Cn3D, and it will show imported structures. To learn more about VAST read the search page.)
Viewing Structures in Cn3D
From http://www.ncbi.nlm.nih.gov/Structure/ type in 1VID and navigate back to the MMDB Structure Summary page. You will view the structure of 1VID in Cn3D by clicking the gray button labeled 'View 3D Structure', and select Best Model, Cn3D, and Display. Cn3D will launch, and display the 1VID structure. From the Cn3D window, choose 'file', then 'save as' and enter 1VID.val . Make sure to always save a displayed Cn3D file - it doesn't 'save' itself anywhere.
Take a moment to also save the 1VID display as 1VID.png You can use this file in a Web document, and also convert it to GIF and JPEG formats using simple image editing programs. The PNG format (Portable Network Graphics) is an 'open source' version of CompuServe GIF.
From the Structure Summary page you can also explore the PDB entry to 1VID, which will allow you to download native PDB files, as well as view some very nice images. You can also navigate 'in circles' (something NCBI is famous for) by following the link to the Entrez Structure Summary. From there, follow the 1VID link back to the 1VID entry. You can also look at the 1VID entries in PubMed from this link.
You can navigate to RCSB and the Protein Data Bank by following the link to the 1VID entry. The PDB is located at http://www.rcsb.org/pdb/
From the 1VID entry, look at the links in the upper left hand corner, and follow 'View Structure' and 'Download / Display File' . The View Structure link will allow you to view the 1VID structure in a number of formats, as well as download JPEG images of 1VID in ribbons and cylinders. Try to spend as much time at the PDB (Protein Data Bank) website at http://www.rcsb.org/pdb/ especially 'Protein of the Month'.
Aligning Structures in Cn3D
Aligning structures in Cn3D is straightforward but by no means 'simple'. You'll need to first download two structures to align. We will align COMT (1VID) and Caffeic Acid5-Hydroxyferulic Acid 35-O-Methyltransferase Ferulic Acid Complex (1KYZ), which are structural neighbors. A similar (and more interesting) alignment can be done using hemoglobin (2HHD) and myoglobin (1M6C). Let's first build our library of files to align:
We'll import these structures in Cn3D, then align one identified chain against another, merge them, save them, and view them.
Step-by-Step
From experience, this is not an easy exercise. It takes practice, and a lot of trial and error. Be patient, and repeat sample exercises like the one above, then hemoglobin and myoglobin, which are a better 'problem posing' exercise.
Practice
This lesson is copyrighted using an Educational Common License, and may be used freely without restriction for academic purposes.
Robert D. Cormia
