What does triage say about translation NCS? 

On Thu, Apr 17, 2014 at 4:25 PM, Yarrow Madrona <amadrona@uci.edu> wrote:

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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Paul Adams
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