(Note: These tutorials are meant to provide illustrative examples of how to use the AMBER software suite to carry out simulations that can be run on a simple workstation in a reasonable period of time. They do not necessarily provide the optimal choice of parameters or methods for the particular application area.)
Copyright Ross Walker 2005

Case Study: All Atom Structure Prediction and Folding Simulations
of a Stable Protein (Folding Trp-Cage Peptide)

By Ross Walker

Stage 1: Building the starting structure.

In all the previous tutorials we have either had a crystal structure available to us or the system has been sufficiently small that we can create the structure by hand. In this tutorial the structure is too complex to attempt to draw by hand and we also do not have a pdb available. As such we need a way to build a linear peptide chain. Fortunately LEaP contains such a tool, called the sequence command.

The amino acid sequence for Trpcage as given in the paper is:

NLYIQWLKDGGPSSGRPPPS

This is expressed in the single letter notation. Before we can enter this sequence in LEaP we need to convert it to the standard 3 letter notation which is what the sequence command expects. The following table gives the conversion:

So the sequence can be re-written as:

ASN LEU TYR ILE GLN TRP LEU LYS ASP GLY GLY PRO SER SER GLY ARG PRO PRO PRO SER

This is not the whole story. LEaP does not automatically terminate the two ends of the chain, so we have to specify our N terminus, by prefixing it with an N, and our C terminus with a C. Thus the sequence we will use in LEaP is:

NASN LEU TYR ILE GLN TRP LEU LYS ASP GLY GLY PRO SER SER GLY ARG PRO PRO PRO CSER

This should give us a sensible linear chain structure from which to start our folding simulation. So, lets get cracking. Fire up xleap. We will use the FF99 force field:

    $AMBERHOME/exe/xleap -s -f $AMBERHOME/dat/leap/cmd/leaprc.ff99

(REMEMBER TO TURN NUMLOCK OFF WHEN USING XLEAP)

Then use the sequence command to build our structure (for help on this command type: help sequence). Note: The line below has been split over 3 lines to make it easier to read. In xleap you should enter it as a single line.

    >TC5b = sequence { NASN LEU TYR ILE GLN TRP LEU LYS
                        ASP GLY GLY PRO SER SER GLY ARG
                        PRO PRO PRO CSER }

The structure will be created as TC5b, we can look at it using the edit command.

    >edit TC5b

So, we have a nice linear starting structure. Since there are many steric clashes, we will need to run a short minimisation of this structure before starting molecular dynamics. Lets save this structure as a library file so we can come back to it in a bit. We will also save a pdb file of this structure so we can easily visualise it in VMD if we want to.

    >saveoff TC5b TC5b_linear.lib
    >savepdb TC5b TC5b_linear.pdb

(TC5b_linear.lib, TC5b_linear.pdb)

CLICK HERE TO GO TO SECTION 2

(Note: These tutorials are meant to provide illustrative examples of how to use the AMBER software suite to carry out simulations that can be run on a simple workstation in a reasonable period of time. They do not necessarily provide the optimal choice of parameters or methods for the particular application area.)
Copyright Ross Walker 2005