Defining sequences and molecular systems

So far we have been linking related spectra with anonymous resonances. In the end we will assign these resonances to specific atoms, but first we must specify the molecules within our experimental sample that we may assign to. Creating a list of atoms to which we may assign has two distinct steps. The first is the specification of a molecule, usually with an amino acid sequence, that will act as a template. The second step is to build an assignable molecular system from the templates to give the atoms. The molecular system represents all the assignable molecules that may reside within the NMR sample, these include proteins, nucleic acids and small molecules. For this tutorial we make a mock molecular system with one protein chain and one small molecule. These two steps exist so that we may define a template sequence once, but have the potential to create several distinct polypeptide chains for assignment, for example if we have a homodimer.

Open M: Molecules: Add Sequence. This will open the Molecule: Molecules popup on the {Add Sequence} tab:

"Add Sequence" is a shortcut command that can create both the molecule and molecular system and add a new sequence to it, all in a single operation (it can also add to existing molecules / molecular systems). The kind of molecule you want to create is controlled by the switches at the top. First set "Input Type:" to "3-Letter/Ccp". Then cut and paste the following sequence:

LYS ALA SER SER PRO SER SER LEU THR TYR LYS GLU MET ILE LEU LYS SER MET PRO GLN

LEU ASN ASP GLY LYS GLY SER SER ARG ILE VAL LEU LYS LYS TYR VAL LYS ASP THR TYR 

(On Linux: left mouse to select in the browser and middle mouse to paste; on Windows: ctrl-C / ctrl-V; on OSX: cmd-C / cmd-V (try to paste with ctrl-v if cmd-v doesn't work)) into the Add Sequence text window:

The input sequence does not have to be perfectly formatted. You can click [Tidy] at any time to see how Analysis has interpreted the sequence:

Also, you can switch between one- and three-letter codes after the sequences has been entered. If you look in the Ccp Codes pull-down menu (top right) you will see all of the residue codes that are currently available, many of which are modified amino acids. Now set both "Destination Molecule:" and "Destination Mol System:" to "<New>", and click [Add Sequence!]. You will be prompted for the names of the new molecular system, and chain. When finished the popup automatically switches over to the {Chains} tab, where you can see the result:

Just for fun, we will now add a small molecule to our project. This could be part of the first molecule we made, i.e. linked to the protein in some way, but instead we will make a new molecule. {Add Sequence} is only relevant for linear polymers so we shall do it another way. First go to the {Small Compounds} tab. Begin by choosing [DNA] in the "Mol Type:" pulldown at the top then select a compound in the right hand table to display an idealised structure in the left hand panel. You can rotate the cartoon compound by clicking and dragging with the middle mouse button. Using 'Shift' or mouse wheel while dragging up and down with the middle mouse button will change the zoom level.

With any compound selected set "Destination Molecule:" to "<New>" and click [Add Compound] to enter it into your second molecule template - this will bring you to the {Sequences} tab. Go back to the {Small Compounds} tab and now try adding a non-polymer molecule. Set the "Mol Type:" pulldown to "Other" and select a molecule at random. Some simple molecules (like Zn) are available, but most will give you a popup asking if it is OK to download the molecule description:

If you click [OK] the description will be downloaded (assuming you have an internet connection and write access to the relevant directory), but even if successful most likely you will also get a warning that there are no coordinates available (that is why we started with DNA). So select something like ATP or Zn. Add that compound to another new molecule.

To make molecular system information from your templates, go to the {Chains} tab. Set "Mol System:" for the new chain' to "<New>", select one of your small molecules as the template, and click [Make Chain From Template]. Then set a new template, keeping the setting for the Mol System, and click [Make Chain From Template] again. You are done creating molecules now; try to look in the various tabs and see what information is available.

To view the new polymer chain have a look at M: Molecules: Atom Browser. You can scroll through the sequence and by toggling the [H], [N] etc. you can display all of the assignable sets of atoms.

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