CCP4i: Graphical User Interface
	Tutorial 6

Refinement

Before you start

Make sure the directory "ccp4itut" is in place
(see Introduction).

Check the presence of the coordinate file rnase.pdb, the reflection files rnase25.mtz and rnase18.mtz in the directory rnase_exam, and the file protin-ccp4itut.def in the directory $CCP4I_TOP/test_data.
Use your experience from Tutorial 1 - Looking at files for this.
If the MTZ files are not there, but only the ascii equivalent .na4 files, create the appropriate MTZ files yourself through Tutorial 1 - Preparing data.

Please note that this tutorial refers to refinement with Refmac before version5.0. This can only be run through the Program List. A tutorial for refinement with Refmac5 (now the preferred option) is available elsewhere.

Protin

Protin is used to set up the restraints for the molecule; these will be used in all subsequent refinement. The Protin program is run before Refmac to generate two temporary files which specify the restaints for Refmac. You must enter some information about your protein but generally you will only need to do this once for each structure, and the information will be saved and used whenever you run Refmac. One day, this will be fully automatic and Protin will not need to be used anymore.

This part of Tutorial 6 is mainly for your information. As you start refinement of your own project, you will have to set up Protin Restraints. When you have provided all the information specific to your own project, save this to a protin.def file in an appropriate place (probably the CCP4_DATABASE directory connected to your 'real' Project Directory). If you already know how to set up Protin Restraints, you can go straight to the Refmac tutorial.

Select Protin Restraints
on the left, in the Refinement module (you may have to select this module first). This opens the Task Window for Protin.
Click on the pop-up menu Save or Restore and select
Restore from File > Select File Name.
From the directory $CCP4I_top/test_data, select protin-ccp4itut.def.

Do not worry about the jumping of windows. Whenever only a shape of a window comes up, position it on the screen where you want it (as before).

The Protin Restraints Task Window is now filled with information needed by subsequent refinement programs:

• At the top, in the Files folder (see also Introduction - Task Window Format), it shows the coordinate file this refinement run will be concerned with, and the dictionary file that will be used.
• In the General Options folder you can specify the amount of information that will be written to the log file. 'Few' should be sufficient.
• Protin is concerned with Chain Types. RNAse SA has two identical protein chains, which will be designated chain type 1, with residues 1 to 96 but no special groups. It also has a sulphate ion, which will be chain type 2 of constitution 'non-protein', and water molecules, which will be chain type 3 of constitution 'non-protein'.
• There are IntRA-chain disulphides in the protein chains, from residue 7 to residue 96. The folder is closed by default, because not all proteins contain intra-chain disulphides.
  Open the folder
  by clicking on the small grey square on the right hand side, to see the description of this disulphide.
• There are no special IntRA-chain distance restraints.
• In the Chain Definitions folder, the actual RNAse chains are linked to the Chain Types. RNAse SA has four chains:

  A and B

  which are the protein chains, and therefore Chain Type 1

  U

  which is the sulphate ion, and therefore Chain Type 2

  W

  which contains the water molecules, and is therefore Chain Type 3

  Since all four chains start with 'residue' 1, the offset is 0 for all of them.
• The remaining three folders (IntER-chain disulphides, IntER-chain distance restraints and Non-crystallographic symmetry restraints) are not used in this simple tutorial.

Close
the Protin Restraints Task Window.

More information

For more information on the Protin program, see PROTIN.

The Interface documentation on features from this tutorial can be found at: