Two possible points. Could there be a problem with twinning, or
spacegroup? The Rfactors seem rather high..
You dont say whether you have a non-crystallographic translation, but it
is faintly possible with 2 molecules that the SG is actually C222 ..
Or that twinning could be present - look at the ctruncate plots - espec
the L test - for infdications..
But re the TLS. You need to set the overall B to the wilson value more
or less. All B factors after REFMAC are relative to that initial starting B
If you TLSANL you should get back sensible B values in the PDB slot, but
of course any which were restricted to the default value of " will be
corruppted..
Eleanor
Christophe Wirth wrote:
Dear CCP Community,
We have processed protein scattering data in space group C2221 to a
resolution of 2.4 Å. The structure shows two protein molecules (chain A
and B) in the asymmetric unit, related by a local two-fold symmetry
axis. Initial rigid body refinement and subsequent full refinement of
isotropic atomic B-factors (including all water and ligand molecules;
using also non-crystallographic symmetry restraints) with the program
REFMAC5 yielded R and Rfree values of 32.1and 33.9, respectively.
In the next step, after setting the B-factor to 30 Å^2, we carried out 6
cycles of TLS refinement followed by 10 cycles of isotropic atomic
B-factor refinement (two TLS-bodies were defined: the two
symmetry-related streptavidin chains A and B in the asymmetric unit). As
expected, the R and Rfree values droped down by 1-2% to 29.6 and 32.5%,
respectively. Checking the residual atomic B-factors their global
average had changed from 13 Å^2 (without TLS refinement) to 2 Å^2, the
lower B-factor boundary. In fact, all atoms in the .pdb file showed
B-values of 2 Å^2.
Interestingly, when we previously processed the same molecule in C2 we
did not have this TLS problem.
Does anybody has an explanation why after TLS refinement, individual
B-factor refinement is failing? Is this due to wrongly refined (i.e. too
large) TLS parameters? Has this been observed before? Any suggestions to
solve the problem?
Many Thanks,
Christophe
