All files for this example are on
hermione in directory:
In this case, the molecular replacement
is done using MOLREP (a program
within CCP4), and refined using anneal.inp in CNS.
The structure of L9 mutant N27C has
2 molecules in the asymmetric unit. This is
different from L9 wild-type, which has one molecule in the asymmetric unit,
crystallized in a different space group.
The unit cell dimensions of L9 N27C are:
a = 131.7
b = 37.5
c = 101.7
space group = P212121
(compare this with wild-type L9,
which is a much smaller unit cell of
a = 135.3, b= 37.3, c = 49.0)
To get started, two things are needed:
1) A search model
for the structure. In this case, L9_mr_model.pdb is used.
This model contains about 2/3 of the L9 wild-type structure.
2) X-ray data
for mutant L9 N27C. This is called "L9_n27c.mtz", and is in
CCP4 format. The data goes to about 2.9 A resolution.
Step 1: Do Molecular replacement
The script for this is a file called "molrep_n27c.sh" :
molrep HKLIN L9_n27c.mtz \
LABIN F=Fnat SIGF=Snat
#NP 10 - number of peaks of rotation function to be checked.
# Maximum: 50. Default: 10
#NPT 20 - number of peaks of translation function to be checked.
# Maximum: 50. Default: 20
#NMON 1 - number of monomers. Program will try to create full model
The output from molrep is a PDB file
with the chains of both molecules that
are in the asymmetric unit. It is called "molrep.pdb".
Step 2: Refine the L9 N27C structure using CNS:
First must convert x-ray data to CNS format, using "mtz2cns.sh" script:
hklin L9_n27c.mtz \
hklout n27c.cns \
labin FP=Fnat SIGFP=Snat
Next, must add free-R flag to data, using "make_cv.inp".
Next, must generate ".mtf" file and CNS-format PDB file using "gen_dimer.inp".
Next, do annealing to refine structure using "anneal.inp".
Finally, calcuate map using "model_map.inp".
Here is a view of the map, as a JPEG file.
All files for this example are on hermione in directory: