The solvent building
procedure within ARP/wARP Version 7.0 has
only marginally changed since the previous release. Within this module
restrained reciprocal space refinement is carried out with REFMAC while
ARP/wARP is performing automatic adjustment of the solvent structure.
Resolution of the data should be 2.5 or higher. The output is the protein
model with the solvent molecules transformed with symmetry operations to lie
around the protein.
The ARP/wARP solvent building module requires
the X-ray data (in MTZ format) and the protein model (in PDB format) without
solvent or with a partial solvent model.
o
MTZ
in X-ray data
in the MTZ format containing structure factor amplitudes and their standard
deviations.
o
Fobs
Sigma If the
MTZ column labels for structure factor amplitudes and their standard deviations
have obvious names, they will be recognised automatically. Otherwise please use
the scrolling button, navigate to List All Labels and chose appropriate ones.
o
Starting
model in
Provide the PDB file with coordinates of the protein only. If the file already
contains some solvent sites, these will be updated during the iterative solvent
building.
o
Output
model Provide
the name of the file where output PDB of the protein with the built solvent
will be written to.
There are a number of options that can be added
either in the main GUI panel or under the Parameters section. You normally should not
need to worry about these. A brief description is given below.
o
ARP/REFMAC
refinement cycles By default 20 cycles will be carried out. Please monitor R factor / R
free for convergence.
o
Free
R flag It is
advantageous to use R free flag for solvent building. You can chose to use
R-free, this will cause additional options to appear within the section Refmac
parameters. The default is not to use R free.
o
Add
atoms This
follows by two numbers defining the threshold (in density sigmas above the
mean) for addition and removal of solvent atoms. The defaults are 3.4 and 1.0,
respectively, which should work for most cases.
o
Disable
Wilson plot statistics check The current Wilson plot checking routine is probably too stringent. You may disable the check and the
warnings if you are sure that the X-ray data is of high quality.
o
Cycles
of refinement in each Refmac run Refmac is invoked to refine the hybrid model
before the density maps are computed. The default is 1 cycle and there is
usually no need to change this.
o
Matrix
weight for Xray / Geometry The default is automatic weighting. This proved to work well and,
probably, there is no need to change this parameter.
o
Scaling
model The
default is to use solvent correction for scaling low angle part of the X-ray
data. You can turn this off (chose simple solvent correct) if your low angle data are
missing (e.g. your data have about 8 low resolution cutoff) or they suffer
from missing overloaded reflections.
o
Scaling
B factor The
default is to use anisotropic B factor for scaling the X-ray data. You can turn this off (chose isotropic scaling B factor) if your data are
systematically incomplete (e.g. a cone is missing in reciprocal space).
o
Data
with free R label
This parameter appears if the free R flag is chosen for refinement of the
protein part of the model. Here you can provide a column label for the free R
flag.
o
Scaling
and sigmaa calculations This parameter appears if the free R flag is chosen for refinement of
the protein part of the model. The scaling and calculation of sA coefficients by Refmac map can be
computed on the bases of the free reflections (this is the default) or using
all reflections.
o
TLS
refinement The
default is not to do a TLS refinement of a hybrid model.
o
Input
a user-defined library file If you already have a Refmac-style cif library for, e.g. your already
present ligand, you can input it here.
o
Space
group, Cell,
ARP/wARP asymmetric unit, Wilson B factor and Solvent content are derived automatically from the
MTZ and the PDB files, displayed for information only and cannot be changed.
However, you may want to check whether their values conform to your
expectations.
o
Resolution By default all reflections present
in the MTZ file will be used. You can check the box (Use reflections
between) and
then narrow the range if you are aware of certain deficiencies of your data.
o
Refinement
with refmac The
R factor (and R free if requested) are printed after refinement of the protein
with Refmac. Check that the value of the R factor is decreasing upon solvent
building.
o
Job
termination The
statement Task completed successfully indicates that the job is finished with no
error. An error statement
QUITTING ARP/wARP module stopped with an error
message: name_of_the_programme indicated that one of the modules of the task
has terminated with an error message. Please refer to the specified log file.
The script file auto_solvent.sh in the $warpbin directory allows you to run the
solvent building as a single-line command without the use of the GUI. The use
of auto_solvent.sh is fairly simple. The script prints out help information if it is
invoked without arguments.
Required keywords are: datafile (followed by the mtz-file name with
the full path) and protein (followed by the pdb-file name of the protein model with the full
path).
Optional keywords include: workdir (followed by the full path to the
working directory), solventfileout (followed by the name of the PDB file where
the output will be written), fp (followed by the fp label), sigfp (followed by the sigfp label) and freer (followed by the Rfree label). The
defaults for the first two are FP and SIGFP, respectively. Alternatively, if
the mtz file contains only one column for structure factor amplitudes and only
one column for their standard deviations, these will be taken. The number of
cycles (default is 20) can be changed with keyword restrcyc. The user-defined library and the
tls-tensor for Refmac can be supplied by using the keywords extralibrary and tlsin.
Example call (assumed to be
started from workdir where test data should reside):
$warpbin/auto_solvent.sh
\
datafile {mtzfile}
\
protein {starting_PDB_file}
\
[workdir {FULLPATH_WORKING_DIRECTORY}]
\
[solventfileout
{output_PDB_file}]
\
[fp {fp_label}] [sigfp {sigfp_label}] [freer {freer_label}] \
[restrcyc {number_of_cycles}]
\
[extralibrary {user_defined_library_for_Refmac5}] \
[tlsin {fixed pre-refined TLS tensors from Refmac5}]
The script will then create a directory in the workdir whose name will be printed and
where a parameter file will be created. The log files and additional output
files as well as the building results can be found in the directory created by auto_solvent.sh.