When you store files you should keep them in "subdirectories" below your home directory. To make a MacroModel directory for yourself, in a winterm or unix shell, while still in your home directory, type:
You should also make a special directory to keep ".pdb" files throughout the semester; make such a directory below your home directory (
mkdir pdbfiles).cd mmod or cd ~/mmod
The squiggle or tilde (~) is a shorthand
symbol for the full path for your home directory. The first command involved
a relative path name and so will only work if you started in your home
directory; the second is an absolute path name and so would work regardless
of where you start from.
Enter Terminal Type: ===> enter the number
2 here.
For every button or command in the MacroModelinterface, the left mouse button activates the command (often "toggles the command off and on") while the right mouse button brings the help information for that command to the screen. This help facility will be very useful for you throughout the exercise.
To become familiar with the MacroModel interface, have a brief look at An Introduction to the MacroModel Molecular Modeling System
Read and view the PDB files a simple
protein, BPTI (1aal.pdb) using Directions
for Viewing Proteins in MacroModel. Some pdb files have been downloaded
into a central directory,
/usr/local/chemistry/MacroModel_Spring97/pdb_files
. We will use these files at the beginning
to practice visualization; since they are in a central directory you can
only read them but not edit or write new ones. Alternatively, if you prefer, you can download your own files.
Build a "capped" alanine by using the follwing sequence of commands:
There are also sidechain (C beta) - mainchain
conflicts. Set phi to +120 and/or psi to -120 to identify why these regions
are poorly occupied.
Build a short poly alanine fragment in the
following conformations: helical (alpha, pi and 3/10), parallel beta and antiparallel beta sheet, using
the
To summarize what you saw, draw a cartoon of the Ramachandran plot, and
identify these 5 basic "unoccupied" regions. Sketch the structure corresponding
to each in a flat or a Newman projection and comment on why it is unoccupied.
Identify the regions of the 5 secondary structure elements mentioned above also.
There are also sidechain torsional preferences, for amino acids with gamma sidechain carbons. Chi 1 is the torsional angle associated with the Calpha - Cbeta bond.
Build a capped valine and a capped tyrosine to examine these preferences. Which is preferred, trans, gauche+ or gauche -?
The pdb file consists of the following parts:
Remember that the discussion of the
cell dimensions, space group possible problems and issues with the structure,
definition of the secondary structure regions, etc. are all also discussed
in the "other links" page of the pdb file, for example
PDBREPORT ("what_check" output) or "PROCHECK" output in the PDBsum.
You might want to compare the headers to these outputs.....
For more details about the pdb file format
see the endless Guide
to PDB files
Look at typical backbone torsional angles
(phi and psi) and typical backbone hydrogen bonding lengths in the
following segments. Compare your values to those that were heavily populated
in the statistical Ramachandran plots linked to the 3DB browser for all-helical
proteins.
You should also look at the links from
the pdb file,
'PDBsum" -> "PROMOTIF summary" -> "Diagram" and
"Motif Description" for the regions of the proteins that you studied,
especially the backbone dihedral angles.
Look at this link .
Go To Top Course Page
Considering Torsional Preferences of the Backbone
and Sidechain: Understanding the Qualitative Features of the Ramachandran
Plot
Before we cut the fragments out of pdb files, we will have a look at some
sections of
"idealized" helces and sheets and also some amino acids that are
deliberately built
into particular backbone torsional states. The objective
is to try to understand
why there are strong preferences in torsional states and whether helices and
sheets are in preferred states. Certain regions of the Ramachandran plot are
rarely populated : for example (phi,psi) = (0,180),(0,0) and (180,0) are
not populated while (180,180) is. (Review your reading on the
Ramachandran plot
from Creighton's book, from the Birkbeck course, or from the Netscape Visualization lesson to refresh your memory on which portions of the plot are occupied and which are unoccupied.) The underlying reasons for certain conformations being unallowed are essentially steric and can be
seen by building fragments and fixing them to these conformational
regions.
PDB files and Formats
In this section of the exercise you will download
a pdbfile and "excise" a particular motif from it for more careful study.
Since you will eventually be downloading lots of files, you will
benefit from organizing your files into a set of "directories" and it is
best to make these directories at the outset by following these directions
for
Making, Looking at and Moving Around in Directories and Downloading Files.
Download the pdbfile for Triosephosphate Isomerase "TIM".
Download 1mbo (myoglobin)
1hbq (retinol binding protein), 2cna (concanavalin a)
and 1cbs (retinol binding protein) into your ~/pdbfiles directory.
Look for all of these sections in the TIM
file you downloaded, using the unix command
In this section you find literature references
for the work that led to this structure (for example you can find details
on the sample preparation). Normally there are remarks about the resolution,
the "R factor" and about the method used to refine the structure. Finally
the authors will often present some remarks about interesting or problematic
sections of the structure. It is often useful to read this section.....
The EMACS Editor:
To become familiar with EMACS have a look
at an Introduction
to EMACS.
Cutting Motifs from Proteins and Measuring Bonds and Angles:
jot (and return)
in a winterm or unix shell). Alternatively you
can make your list in the
EMACS editor or the Netscape editor.
Alpha Helices
Look at N terminal capping hydrogen bonds
in the same segments and identify the residue involved.
Look at the conformation of Proline in
Helices. Proline is often said to deform helices because of the constraints
imposed by the ring structure, and because of its inability to participate
in backbone hydrogen bonds as the donor. In addition, there are additional
steric clashes between its sidechain carbons and the backbone atoms of
the preceding residue; can you identify what conflict this is and which
conformations would be unallowed especially for proline? Measure the Ramachandran
Phi and Psi values around proline, especially the residue preceding the
proline.
Find a bent helix and comment
on the deformations in the backbone torsional (dihedral) angles for bent
helices. You could locate one yourself using your textbook or using the
3DB browser and RasMol, or you can use one of the long coiled-coil in the
serine tRNA synthetase that you looked at previously
(in the netscape exercise),
which is obviously bent (this is residues 25-100:A).
Beta Sheets
The retinol binding protein structure exhibits
the typically right handed twisted sheets. Measure
the phi and psi dihedral values for a few residues and then for
the serines. Compare the values to the regions in which left handed and
right handed twists are expected. The region for right handed twisted sheets
is just above the line phi = -psi (e.g. phi=-120, psi=140); the region
for left handed is just below that line (e.g. phi =-120, psi =100). It
may be worthwhile to build some structures that are uniformly in each region,
so that you can visualize these twists. What differences do you see
between the parallel and the pantiparallel structures in terms of
backbone torsinal states?
Beta Turns
Concanavalin A (jack bean) "2cna" contains
many beta turns and hairpins. Measure the phi and psi values for
the 2 residues (central 2) in the turn and compare to the Birkbeck plots.
Measure the phi and psi values for the 2 residues (central 2) in the turn
and compare to the Birkbeck plots. Note that the PDBsum page (in "other links")
shows the
location of beta turns in cartoon format, so you could in principle pick
beta turns from any protein in the PDB that has one. See Birkbeck
College Course for nomenclature.
Gamma Turns
Use the PDBsum (in other links) "linear" display of secondary structure for TIM and hemoglobin to identify a gamma turn and measure its backbone
dihedral angles in MacroModel.
Find beta bulges in the proteins analyzed
above. You can use the PDBsum analysis or the PROMOTIF summary, as you did
in the first exercise. Discuss their backbone dihedral angles.
Sidechain Dihedral Angle Preferences
Select a few tyrosine and valine residues
in the proteins discussed above. Measure their chi 1 sidechain dihedral
angles. Discuss the results in terms of the trans, gauche+ and gauche -
rotomer positions: which are expected to be occupied, and which are occupied?
PROCHECK: Checking
Your Results and Looking at Many Other Local Structural Features
To check your tables of results, use PROCHECK.
This program will generate lots of output files, so you should make a directory
called "PROCHECK" below your home directory. Change to your PROCHECK directory
and type,
"Procheck ~/pdbfiles/filename.pdb #"
where ~/pdbfiles/filename.pdb
is the name of some pdbfile you previously downloaded into your
~/pdbfiles/
directory, and "#" is the resolution
of the structure (in Angstroms, e.g. 2.0). The output of this program will be in a series
of newly generated files including "filename.sum" and several files called
"filename_ #.ps" where # are numbers that run from 01 to 10 or so.
These postscript (.ps) files are graphics files containing plots
of the dihedral angles for your protein, including phi,psi plots and sidechain
dihedral plots. To view these files type "xpsview filename
*.ps" .
You should see the Adobe ShowPS viewer appear with the first page
(filename01.ps)
of your output. In the pull-down menus select from "view" the selection
"shortcutpanel" so that you can scroll through the pages. You can
compare the entries on these plots these to the dihedral angle values you
got using MacroModel. As you can see, this is a very convenient tool for
obtaining backbone and sidechain dihedral angles in a glance.
A Discussion of these Structures
We have prepared several fragments and
a discussion of the structures .
Bringing Other File Formats into MacroModel or RasMol
Frequently coordinate files have formats other
than the pdb format you learned about today. For example, coordinate files
for entries in the cambridge database, which you will use next week,
can be written
using the "Quest" software,
but they are not pdb formats and in fact they cannot be read by the software
we are learning (RasMol, MacroModel, spartan, insight).
A program (freeware) called "Babel" can be used to interconvert
common formats.
Your Homework
Go To Schedule Page and List of Exercises