Dear all,

     While the thread on "Another MR pi(t)fall" is still lukewarm, and
the discussion it triggered hopefully still present in readers' minds,
I would like to bring another puzzling entry to the BB's attention. 

     When reviewing on Monday the weekend's BUSTER runs on the last
batch of PDB depositions, Andrew Sharff (here) noticed that entry 5gnn
had been flagged as giving much larger R-values when re-refined with 
BUSTER (0.3590/0.3880) than the deposited ones (0.2210/0.2500). This
led us to carry out some investigation of that entry.

     The deposited coordinates were flagged by BUSTER as having 4602
bond-length violations, the worst being 205.8 sigmas, and other wild
outliers. The initial Molprobity analysis gave a clash score of near
100, placing it in the 0-th percentile. The PDB validation report is
dominantly red and ochre, with only a few wisps of green. 

     Examining the model and map with Coot showed "waters, waters
everywhere", disconnected density, and molecules separated by large
layers of water. The PDB header lists hundreds of water molecules in
REMARK 525 records that are further than 5.0 Angs from the nearest
chain, some of them up to 15 Angs away.

     The cartoons on the NCBI server at 

http://www.ncbi.nlm.nih.gov/Structure/mmdb/mmdbsrv.cgi?uid=142582&dps=1

show random coils threaded up and down through beta-strands, and the
one on the RCSB PDB site at 

       http://www.rcsb.org/pdb/explore.do?structureId=5GNN

also shows mostly random coil, with only very few and very short
segments of secondary structure.

     In reciprocal space, an oddness of a different kind is that if
one looks at the mtz file, the amplitudes and their sigmas are on a
very small scale. However the STARANISO display shows a smooth and
plausible distribution of I/sig(I) to the full nominal resolution
limit of 1.6A.

     Looking at the publication associated with this entry
     
             http://www.ncbi.nlm.nih.gov/pubmed/27492925

indicates that the structure was solved by MR from a model obtained
from a structure prediction server (I-TASSER). No further details are
given, even in the Supplemental Material. Table 1 does report a
MolProbity clash score of 103.59, as well as 10% Ramachandran outliers
and 25.51% rotamer outliers. It also contains a mention of a twinning
operator -h, -k, l with a twinning fraction of 0.5, although there is
no mention of it in the text nor in the PDB file.

     I will follow my own advice and resist the temptation of calling
this "the end of civilisation as we know it", but this is startling.
Perhaps we have over-advertised to the non-experts the few successes
of structure prediction programs as reliable sources of MR models and
thus created unwarranted optimism, besides the usual exaggeration of
the degree to which X-ray crystallography has become a push-button
commodity that can deliver results to untrained users. What is also
disconcerting is that the abundant alarm bells that rang along the way
(the MolProbity clash score and geometry reports, the contents of the
PDB validation report, and simple common sense when examining electron
density and model) failed to make anyone involved along the way take
notice that there was something seriously wrong.

     This case seems to bring to the forefront even more vividly than
4nl6 and 4nl7 some collective issues that we face. Here the problem is
not one of contamination of a protein prep resulting in crystals of
"the wrong protein": there is also a more diffuse contamination by
deficiencies of judgement, expertise and vigilance at several
consecutive stages, including refereeing and publication.

     Validation is a hot topic at the moment, and this may serve as a
concrete example that some joined-up thinking and action is indeed a 
matter of urgency, and that extreme scenarios of things going wrong do
not exist solely in the imaginations of obsessive-compulsive/paranoid
validators.

     I am grateful to several colleagues for correspondance and
discussions on the matters touched upon on this message.


     With best wishes,
     
          Gerard

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     * Gerard Bricogne                     [log in to unmask]  *
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