MSc Toxicology Introduction to Structural Bioinformatics Course
Introduction to Structural Bioinformatics Course
In this exercise we will look at the steps need to
produce a model of the structure of human platelet sulfotransferase SULT1A3
with its substrate dopamine. These notes can be found by following
the link from Oliver Smart's home page
www.bip.bham.ac.uk/osmart
Introduction
Setting the scene a talk:
From sequence to structure to function (Maybe!): A quick guide to structural modelling.
The exercise
We are going to attempt to do a great deal in a short time.
For each part of the exercise - follow the leader - you will be
shown what to do on the projector - the do it yourselves.
If you want to be "signed off" as having completed this course
then you need to print out and complete the
assessment questions sheet and
hand them in. When you see the happy little sign:
in the margin, this means that there is a question .
- Logging on and out from an sgi, starting netscape (minimal unix).
Follow this this link if confused.
- Background: The sulfotranserases are a family of enzymes that have a physiological
role in the sulfonation of a wide range of biologically important molecules,
including steroids. They are also intimately involved in toxicological behaviour of
of endocrine disruptors most notably xenoestrogens. Students should familarize
themselves with the area by reading the introduction to
Harris R. M. et al.
J. Biol. Chem. (2000) 275:159-166. The first part of this exercise is
to find this reference - no need to rush down to the Barnes simply find the
J. Biol. Chem. web site starting at the
University main page (index, i, information services,
A-Z of Services, E, Electronic Journals, Electronic Journals, A-Z lists, I-J,
J, Journal of B., Journal of Biological Chemistry on line, Search the JBC,
Harris, R)
.
- Finding a sequence. We are going to use the "SWISS-PROT" database
from EBI (European Bioinformatics Institute). Find it from
the following link www.ebi.ac.uk
Then search for "human phenol sulfotransferase". Find the
entries from SULT1A1, SULT1A2, SULT1A3. Find the required information
to answer questions. Also download the sequence of sult1a1 in pir format.
- Finding homologous sequences by running a blast.
Run a blastp of SULT1A3.
- Finding a protein 3D structure.
Go back to the ebi main page www.ebi.ac.uk
and find the EBI Macromolecular Structure Database.
Use the 3DB browser to search
for sulfotransferases. Identify the pdb codes for
(a) the estrogen sulfotransferase
with 17-Beta Estradiol bound
and (b) human SULT1A3.
We want to download monomeric structures of each
but the estrogen sulfotransferase crystal contains
two individual copies of the protein in the crystal.
So for estrogen sulfotransferase do not use
the PDB entry "save to Disk" option instead follow
"Likely Quarternary Molecular Structure file(s)" option and download
a single chain.. .
- Displaying the protein structure with sybyl. You will be shown
how to start sybyl
and to read the pdb structure for estrogen sulfotransferase.
Display the molecule in Calpha mode and practise rotating in.
The cofactor and PAP molecule will not be correctly connected -
ask the demonstrator how to achive this.
Use this link to find out how.
- Displaying two protein structures.
Read in the pdb file for estrogen sulfotransferase that you downloaded
in (5) into molecule area M2. Also display this as Calpha only.
Does it seem similar to the sult1a3? Try using the
Align structures using homology feature Colour the two
molecules differently. Use the
Display Area Options widget
to get the
Display Area Options widget
dialogue box:
and click on the D1 and D2 buttons to turn the
proteins' display on and off.
Assess how the proteins are similar and how they differ.
- Making a model for dopamine.
The demonstrator will show you how to use sybyl's "Sketch molecule" facility
to make a model of dopamine.
Or use this link to download a
sybyl .mol2 of "one I prepared earlier" Use "shift-left click
to force netscape to save to disk and then the File Read option
in sybyl to put into a new molecular area.
- Homology modelling
There is not enough time to actually build a homology model of sult1a1
in this exercise. But by using swissmodel it is possible
to get a model for the enzyme. Here it is in sybyl format
sult1a3 full model in mol2 format
- Docking.
Dock you dopamine model into the the sult1a3 full model using est as a guide.
Your demonstrator will show you how to the "Analyze" fit atoms
to place your dopamine model onto the estradiol from EST.
- Producing a pretty picture
Use the "assign secondary structure" option (in biopolymer) in sybyl to produce a
nice schematic of the protein. Use the mix rendering to
get the dopamine represented as solid spheres. Then use "print screen
capture output" (file menu) to get the picture into a graphics file
(use create and close). You should get something like:
To print the file start
imgworks from a separate unix shell and select print.
