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Hello,
I'm almost done refining a protein-DNA complex at 3A, but when I apply TLS (Refmac+TLSanl to include the ANISOU component) the B factors increase from 50-60 to 100-110 A2. The crystal is a bit special since it has 80% of solvent and the Wilson B-factor is 87 A2. When I compare my Bfactors with other structures of similar resolution using Phenix Validation, my data is clearly an outlier.
To perform the TLS refinement I have generated .tlsin files for 2,4 and 6 groups with the TLSmd server and used them in refmac jobs consisting of 20 cycles of TLS refinement+50 cycles of restrained refinement. This way the R-factor and R-free values have drop 1-2% (before it was 22.3/24.1). The three different options suffer from the same problem regarding high B-factors after TLSanl. Am I doing something wrong during the TLS refinement? What can I do to preserve the original Bfactors?
Thanks in advance,
Carles Palanca.