Following program installation, RSRef2000 needs to be set up in each directory in which O is to be run.
The setup procedure creates a new User Menu in O. If the user menu has already been customized, its contents should be saved, and following setup, read into the file $RSdir/omacros/rsrefmenu.o. In this way both the options required by RSRef2000, and the user's will be displayed.
Setup initializes a number of files. Edited / customized values may be lost if setup is run again in the same directory.
Run @RSdir/omacros/rsrefSetup.macro as a macro from within O. (This option is not available under Windows.) RSdir is the environmental variable that defines the full path to the RSRef2000 installation. Such environmental variables may not be accessible within the O program, so you will likely have to type the full path explicitly within O. Example: @/ser/c/users/RSRef2000/omacros/rsrefSetup.macro
Copy $RSdir/pscripts/rsrefGUI.py to the directory in which O is to be run. Run rsrefGUI.py and in the resulting session of RSRef GUI editor click “Prepare for ‘O’” (this should only be done once per directory, even if repeating the setup). Then type @rsrefSetup.macro in the O window.
Name of the molecule you want to refine ("mol" name in O)
Name of the RSRef2000 parameter file with its absolute path. Note that this parameter file is changed and overwritten during refinement, so should be backed-up periodically to a file of a different name.
Under Windows, it is compulsory to be running rsrefGUI.py (that controls the graphics user interface), and it should be started once per O-session. Under Unix, it is recommended, but not necessary.
Aside from the selection of residues, this is the last time that refinement parameters may be changed under Unix. They may be edited with the graphics user interface (Windows or Unix) or through text-editing of the parameter file (Unix only). Note that saved parameters may be read from a file. Note also that the latest parameters will continue to be used until changed again, but need to be saved explicitly for use in another session.
If the setup has completed correctly, the O window should contain an RSRef menu with 4 options that (optionally) add residue selections to the parameter file:
to pick the first and the last residues of a zone of residues to be refined.
Note that for both zone of residues and spherical region (below) the following applies: If crystallographic or non-crystallographic symmetry is present, the symmetry relationships between the refined region of the protomer and its copies will be retained. Only atoms of the selected region will be refined, while the atoms that fall within margin (determined in the parameter file) will be fixed during refinement. If atoms of the symmetry related subunit and not the 1st protomer are chosen to be refined, the coordinates of the 1st protomer will be changed in order to retain the symmetry. If more than one equivalent of an atom is chosen to be refined at once (for example, when spherical region is defined) the symmetry relationship between them will be retained as well.
to pick an atom at the center of a sphere to be refined. The radius and the width of a margin containing non-refining atoms contributing to the energy calculation are entered from the keyboard.
Like RSRefSph, except that the coordinates of the center are entered by keyboard, not by atom-picking.
removes the object RSRef (when rejecting the changes made by refinement).
Under Unix, the macro will start refinement directly.
Under Windows, refinement must be started explicitly within the graphics user interface by clicking "Run from 'O'".
Under Windows, you will have to tell O when the refinement is complete by typing yes on the O command line, or clicking yes in the O window. (This may need repeating, but do so only until the question “Refinement completed, accept new coordinates Yes/No?” appears at the top of the O modeling window and in the command line.
Under both Unix and Windows, a refined partial model will be placed in an O object, RSRef, and can be inspected.
Answer yes or no to the question: “Refinement completed, accept new coordinates Yes/No?”. If yes, the refined region gets incorporated into the initial model. If no, the RSRef object can be cleaned up with Del_RSRef in the Rsref-O menu.
There are three input files for CNS. They may be changed if, for example, user-defined topology files are to be used:
$RSdir/pscripts/generateLOC.inp - input CNS file used for creating .mtf file before actual refinement.
$RSdir/pscripts/realcgLOC.inp – input CNS file used for conjugate gradient refinement.
$RSdir/pscripts/rsrefLOC.inp - input CNS file for simulated annealing refinement.
If both REMOTE and FRONT END machines are used, these files are located on the FRONT END machine. Under Windows, the files are being temporarily copied to the RSRef_Temporary_Fls directory on the REMOTE host where calculations proceed.ANDREI KOROSTELEV / MICHAEL S. CHAPMAN; Last updated 03/19/2002