Embedded Files
Some of my friends have pointed out that Bioinfo guys with a Bio bend are finding this project kind of confusing. So, here's a refresher. I have added some videos that would help you guys to get up and running.
https://www.youtube.com/watch?v=eZfyWdHnzR0
https://www.youtube.com/watch?v=eF40NMhpuGc
https://www.youtube.com/watch?v=uDp7LGZJn6Q
Knowledge about the workings of the following software would help you get up and running in no time:
1. clustal, clustal-x/w/o
2. CN3D
3. pymol
4. ugene
5. modeller
6. hmmer
7. ncbi-blast
Moreover, you should already have a knowledge about the following:
1. homology modelling, protein threading, gene threading, multiple sequence alignment.
2. signalling networks, nf-kb pathway, Mal Cascade, Akt-jun pathway, pellino proteins etc.
Chapter 0
Foreword:
========
We would be performing molecular modelling and drug designing -- first using computer modelling programs (this is called dry lab) and then verifying our results in vitro (that is to say the shit we do on petri dishes. This is called wet lab). Dry lab phase takes around 6-8 months to perform. You'll also learn the basics of wet lab within this period.
Wet lab takes another 6 months. That would require a real laboratory. We'll get to that later.
you are free to engage your friends in this project if you want. However, I dont want to answer everybody's problems. Everybody's doubts need to come through you.
Chapter 1:
Dry lab -- Introduction
===================
This is an overview of everything we would be doing. You are obviously not expected to understand any of this at the moment. This is being given just to present you the complete picture. What we are trying to do goes like this:
1. Zero in at the exact problem causing molecule.
-- NFkB
2. Pick a problem to solve.
(Parkinsons/Diabetes/Cancer/alzheimer's/AIDS)
3. Trace the pathway of the disease (across all its hosts).
[[First pick the disease. Then I'll tell you about the pathway]]
4. Trace the pathway of the molecule related to the diesase.
[[First pick the disease]]
5. Trace all other pathways of the molecule.
http://www.cellsignal.com/common/content/content.jsp?id=pathways-nfkb
6. Retrieve the sequence of both #4 and #5.
(using BLAST/PDB/ENSEMBL)
7. Perform theoretical multiple alignment.
Needleman Wunch Alignment, Chow Fasman Method, Newton Raphson Method, Monte Carlo Simulation.
8. Verify results.
(mostly using Clustal, HMMER and Modeller)
9. Determine motifs.
Using results from #8
10. Perform fine precision molecular modelling.
Using Rasmol/Pymol/Cn3D.
11. Reduce noise.
Using Ramanujan Plots and Heat maps.
12. Refine #10 using #11.
EXERCISES:
-------------
1. Study the structure of NFkB
Go to this site
http://www.rcsb.org/pdb/explore.do?structureId=1svc
Explore the site. You can see the Validation Report as well as Ramachandran Plot here. We'll get to those later. You can click on all the other links and explore.
Download the text file. http://www.rcsb.org/pdb/download/downloadFile.do?fileFormat=pdb&compression=NO&structureId=1SVC
Study the structure of the text file. Then write a java program to that reads the file and displays all the amino acids on the helical part of the structure.
HINT: the java program reads the file line by line and when it reaches the letters 'H','E','L','I','X', (at lines 448-455 in this file) then returns the corresponding aa (Valine through Proline)
Go to this link. http://www.rcsb.org/pdb/explore/jmol.do?structureId=1SVC&bionumber=1
and then view the 3D structure.
repeat the same for this molecule
http://www.rcsb.org/pdb/explore.do?structureId=2v2t
2. Rasmol
Download here. http://www.rasmol.org/software/RasMol_Latest_Windows_Installer.exe
Read the quickstart: https://www.umass.edu/microbio/rasmol/rasquick.htm
Read the manual: ftp://www.bio.umass.edu/pub/shareware/rasmol/distrib/rasmol.hlp
Quick tutorial: https://www.umass.edu/microbio/rasmol/coulson.txt
Video Example: https://www.youtube.com/watch?v=UllnXjct1YE
Load 1SVC.pdb that you downloaded earlier and play.
3. PyMol
Download here: http://sourceforge.net/projects/pymol/files/latest/download
Read quickstart: http://www.pymolwiki.org/index.php/Practical_Pymol_for_Beginners
Read the manual: http://pymol.sourceforge.net/newman/userman.pdf
Quick Tutorial: http://www.bio.ph.ic.ac.uk/~scurry/pdfs/PyMOL_tutorial.pdf
Video Example: https://www.youtube.com/watch?v=1cIE9owcy2s
Load 1SVC.pdb that you downloaded earlier and play.
4. SwissPDB Viewer
Download here: http://spdbv.vital-it.ch/download/binaries/SPDBV_4.10_PC.zip
Read quickstart: http://spdbv.vital-it.ch/main_guide.html
Read manual: http://spdbv.vital-it.ch/Swiss-PdbViewerManualv3.7.pdf
Quick Turorial: http://www.genebee.msu.su/spdbv/text/activsit.htm
Video example: https://www.youtube.com/watch?v=DSHhep_w6pk
Load 1SVC.pdb that you downloaded earlier and play.
5.Understanding the file format.
http://www.rcsb.org/pdb/101/static101.do?p=education_discussion/Looking-at-Structures/intro.html
<< Visit all the links provided at the right under the legend "Looking at structures" >>
Do It Yourself Exercises:
----------------------------
1. Try to understand what is BLAST.
2. Try to learn VMD and UCSF Chimera.
End of Chapter Prologue:
-----------------------------
In this exercise, you got a very rough idea of the workflow. You got to see the pdb file format. There are other file formats, viz., FASTA, Entrez etc. We'll get to them shortly. You got to see the pdb website that provides extensive structure related information. There are other sites such as NCBI, EMBL, UCSC etc that are used for other purposes. We would get to them in the next chapter.
You got a first hand idea of the various tools that would be used. Rasmol is not used much. But it is the simplest of them all, so acts as a very good starting point. Pymol has extensive scripting capabilities. SPDBV chomps mathematical and physical data beautifully, whereas, VMD is used to deal with molecular dynamics.
In the next chapter, we would visit the various websites that would help us provide all data.
In chapter 3, we would study the theory part and then in chapter 4, we would revisit this chapter with real world examples.
At the end of this, you should be expected to be able to do all of the following:
https://www.youtube.com/watch?v=meNEUTn9Atg&list=PLC482A348BCB4401A
https://www.youtube.com/watch?v=rTNvPDUFmoQ&index=3&list=PLC482A348BCB4401A
https://www.youtube.com/watch?v=TTtrk0Ue-Cg&index=4&list=PLC482A348BCB4401A
https://www.youtube.com/watch?v=3BTRVtsmXpw&list=PLC482A348BCB4401A&index=5
https://www.youtube.com/watch?v=8O3qEtH76OA&index=6&list=PLC482A348BCB4401A
https://www.youtube.com/watch?v=hd2YaygJC-w&index=9&list=PLC482A348BCB4401A
https://www.youtube.com/watch?v=TZM8EzHW6MA&index=7&list=PLC482A348BCB4401A
Follow the instructions at System Setup and System Config to set up your workstation.
GIT Config
-------------
<< Get to your repo folder >>
$ cd
$ cd ~/myrepo
$ cd your_project_name
<< First time cloning >>
$ git clone https://github.com/schacon/ticgit
$ git remote add pb https://github.com/paulboone/ticgit
$ git remote -v
<< Subsequent pulls >>
$ git pull origin master
$ git pull ci master
<< if you made some changes but do not want to keep them >>
$ git checkout master
$ git checkout HEAD~2 // for specific heads
<< for a hard reset >>
$ git reset --hard origin/master
<< to view your logs >>
$ git log --all --pretty=format:'%h: %s %d'
<< for interactive session >>
$ gitk --all
<< amend commits >>
$ git commit -m 'initial commit'
$ git add forgotten_file
$ git commit --amend
<< rejecting changes for specific files >>
$ git checkout -- CONTRIBUTING.md
<< to keep the uncommitted changes for later >>
$ git stash save --keep-index --include-untracked
$ git stash list
$ git checkout -b temp
$ git stash apply //if you want to apply your changes
$ git stash pop //will get everything back in place
$ git stash drop //if you want to get rid of your changes
$ git status // some files might remain
$ git checkout -- <<filename>> // if you really want to get rid of all changes
<< After the changes have been made >>
$ git add .
$ git commit -m 'your favorite commit message '
$ git push origin master
$ git push ci master
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