Hi Yarrow,

the problem is that during structure solution, many wrong paths may have to be followed until finally identifying the correct path.

So the general answer to this kind of problem is: in some way, your parameterization of the experiment is wrong or incomplete.

>From what you write, data quality does not seem to be the problem. But: did XDS really integrate _all_ the reflections, or only a subset (say, every second reflection)?

Check out http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/Refinement#what_can_go_wrong_in_refinement.3F

If, after thorough attempts, you fail to find the solution, upload your current model, sequence and raw data frames to a Dropbox folder and post the link here - there may be people who succeed in processing the data nicely, or otherwise can identify the problem based on the data (rather than based on your description only).

HTH,

Kay

Am 18.04.14 01:25, schrieb phenixbb-request@phenix-online.org:

Date: Thu, 17 Apr 2014 16:25:00 -0700
From: Yarrow Madrona<amadrona@uci.edu>
To: PHENIX user mailing list<phenixbb@phenix-online.org>
Subject: [phenixbb] Stalled refinement
Message-ID:
        <CAMHjG6bPE4q1xWidpE2VwFMJ9qLSqjtLRuLMM1ef9vWBHBfZKg@mail.gmail.com>
Content-Type: text/plain; charset="utf-8"

Hello,

I using the latest stable build of phenx.refine (1.8.4) I recently
collected data, processed and obtained an MR solution using phaser. I am
stuck trying to refine with an Rfree sitting at 40%

I really want to know if the high Rfree is due to poor data quality or if
non-crystallographic symmetry involving a near perfect two fold rotation
between the two molecules in the ASU could somehow impede refinement. Stats
and other information is below. Thank you for any help you can give.

-Yarrow


Visually, the quality of the data is marginal at best (streaky/ice rings in
many frames) despite good processing stats from XDS. Processing with mosflm
or HKL2000 managed to index but failed pretty bad in integration and
scaling.

Phaser gave high TFZ scores for 2 molecules in the asu (see below).

Density for a cholesterol like ligand shows up even though not present in
the search model.

MolRep Self rotation shows rotational symmetry.
https://www.dropbox.com/s/2zsajl5o091k50r/CYP142A2-032814_21_rf%20copy.pdf

The 2 molecules in the ASU are related by almost a 2 fold rotation:

Rotation matrix for chain A to chain B:

new_ncs_group
rota_matrix    1.0000    0.0000    0.0000
rota_matrix    0.0000    1.0000    0.0000
rota_matrix    0.0000    0.0000    1.0000
tran_orth     0.0000    0.0000    0.0000

center_orth   15.2016    0.5245   33.7070

rota_matrix   -0.9860   -0.1636   -0.0309
rota_matrix   -0.1659    0.9511    0.2605
rota_matrix   -0.0132    0.2620   -0.9650
tran_orth      34.3310  -24.0033  107.0457

center_orth   15.7607    7.2426   77.7512

RMSD, B onto A = 0.0007 after phaser
RMSD, B onto A = 0.347 after one round of refinement in phenix


Refinement using aniostropically corrected data (ucla web server:
Services.mbi.ucla.edu/anisoscale) did not improve the Rfree in refinement.


Statistics are listed below:

UNIT CELL: 51.487 88.923 89.592 90 97.15 90 P21

RESOLUTION     NUMBER OF REFLECTIONS    COMPLETENESS R-FACTOR  R-FACTOR
COMPARED I/SIGMA   R-meas  CC(1/2)  Anomal  SigAno   Nano
    LIMIT     OBSERVED  UNIQUE  POSSIBLE     OF DATA   observed  expected
                                    Corr

      5.99        8280    1927      2087       92.3%       3.1%      3.3%
   8246   35.09      3.5%    99.8*    20*   0.909    1296
      4.30       14606    3401      3487       97.5%       3.3%      3.5%
  14580   33.37      3.8%    99.9*    11*   0.843    2273
      3.53       17961    4244      4445       95.5%       3.8%      3.9%

  17944   31.11      4.4%    99.8*    -2    0.789    2721
      3.06       21954    5068      5221       97.1%       4.9%      5.1%
  21933   24.81      5.6%    99.7*    -2    0.780    3455
      2.74       25741    5830      5933       98.3%       7.6%      7.6%
  25713   18.88      8.6%    99.5*    -2    0.782    4165
      2.51       27859    6311      6483       97.3%      10.8%     10.8%
  27824   14.06     12.3%    99.1*    -2    0.774    4385
      2.32       31336    6979      7084       98.5%      14.9%     15.3%
  31296   10.49     16.8%    98.5*    -4    0.748    5095
      2.17       32396    7347      7567       97.1%      22.3%     22.7%
  32341    7.46     25.4%    97.3*    -7    0.728    5055
      2.05       32254    7339      8047       91.2%      33.1%     33.5%
  32075    5.06     37.5%    94.8*    -6    0.724    5155
     total      212387   48446     50354       96.2%       7.8%      7.9%
211952   16.57      8.8%    99.7*    -3    0.768   33600

Processing with mosflm or HKL2000 managed to index but failed pretty bad in
integration and scaling.

Phaser:

SOLU SET RFZ=27.5 TFZ=24.2 PAK=0 LLG=1711 RF++ TFZ=64.6 PAK=0 LLG=3610
LLG=4865

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