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On Wednesday 08 February 2006 02:36 am, you wrote:
>
> I am refining a structure against a 2.1 A resolution remote dataset of a
> Se-Met MAD experiment. Since the R/Rfree is stuck at 21/27 % I tried to
> use TLS refinement. This yielded a much better R/Rfree of 19/24.1 but I
> got this 'Problem in MAKE_U_POSITIVE' warning. Analysing the models
> before and after TLS showed the following results:
>
> no ncs, no tls:
> average B (overall)
> chain A: 48.7
> chain B: 45.6
> chain C: 33.9
> chain D: 20.2
>
> R/Rfree 21/27 % (CNS gives nearly the same results)
>
> no ncs, one tls group per protomer:
> average B (overall)
> chain A: 13.1
> chain B: 12.9
> chain C: 14.5
> chain D: 12.9
>
> R/Rfree 19/24.1
How are you calculating these average B's?
Please be aware that after TLS refinement in refmac,
the column of the PDB file reserved for "temperature factor"
actually contains the residual contibution to the total
isotropic B factor. That is, it contains the part not
described by your TLS model. If you really want to
compare average B's from TLS and non-TLS refinement, you
need to use something like the following jiffy script:
#/bin/csh
#
# Usage:
# tls_correct 1XYZ
#
# where 1XYZ is the 4 letter PDB identifier code for a structure
# containing an explicit TLS model description in the header records.
# This script will create an output file 1XYZ_bcor.pdb
# in which the TLS description has been applied to all relevant
# atoms, creating a file with ANISOU descriptions of these atoms
# rather than TLS + Biso.
#
#
tlsextract XYZIN {$1}.pdb TLSOUT {$1}.tlsout
tlsanl XYZIN {$1}.pdb TLSIN {$1}.tlsout XYZOUT {$1}_bcor.pdb << EOF
BRESID true
ISOOUT full
END
EOF
Then look in the "temperature factor" column of the corrected pdb file.
>
> I would like to know if it is legitimate to use tls only on parts of the
> asymetric unit and/or if there are any other things I should look for.
Sure. There is no guarantee that any particular protein molecule,
or piece of a protein molecule, is well-described by a TLS model.
It often is, but not always.
It may also be the case a multi-domain protein that the individual
domains have large TLS components, but the entire protein taken as
a whole is not well described as a single TLS group. In such a case
you can get a dramatic improvement in R and Rfree by breaking the
protein into multiple TLS groups. However, based on the R factors
you report I doubt your structure is an example of this.
Feed your model to the TLSMD server and see what it says about
your individual chains.
http://skuld.bmsc.washington.edu/~tlsmd
> additional informations:
> The structure solution was straight forward. The experimental density
> for all 4 subunits was good.
>
> spacegroup: P2(1)
> cell axis: 90.570 57.020 152.697 90.000 103.352 90.000
>
> wilson B from XSCALE: 33.23
>
> Rsym (overall/last shell): 5 (25)
> reflections: 86204 (7972)
>
> overloads: 143 out of 1343616
>
> As far as I can see the data looks good so far. the Wilson line is ok
> according to what I see in truncate.
> (it didn't pass the arp/warp check, but until now I haven't come across
> a dataset that passed it).
> The dataset doesn't show any sign of radiation damage (checked this
> looking at the xdsstat output.)
>
--
Ethan A Merritt
Biomolecular Structure Center
University of Washington, Seattle 98195-7742
