Hi All,

Thanks for all the replies, I would like to add more information, after 
reindex it to P21212, the cell parameter is a=88.71, b=116.26, c=55.12, the 
molecule is a long rod like head to head dimer with a length of 110Å (55Å 
long for each monomer) and we used the dimer to search the orthorhombic 
data, two solutions were found as shown in the original thread which can be 
both refined to almost the same very good R/Rfree, coot was used to 
generated the symmetry related molecules from both solutions and the two 
solutions had the same packing but translated along the b axis by 31.9Å, 
thus clashed into each other and made a mess in the overlap region, so I 
think they are not crystallographically the same solutions, unless the 
origin of the cell in P21212 can be placed on b-axis arbitrarily, and I 
think Phaser will also prune the crystallographically same solutions.

Interestingly, the most prominent pseudo-translational peak (1/3 of the 
origin peak), has a fractional vector 0.5000  0.2741  0.5000, and the 
fraction on b axis = 0.2741*116.26 = 31.8755Å, and that is exactly how long 
the two solutions translated along the b-axis, I don't know what that 
means, does these information verify this is the second case Eleanor 
suggested? if so should I keep the messy overlapped region and set the rest 
0 occupancy to check the density?

Thanks for all the suggestions so far!

Xiaoli


>  From the Pattersn peak  it seems very likely that you have two molecules
> in the asymmetric unit seperated by the very vector that seperates your
> two MR solutions, and both MR solutions are correct?
>
> Or is that not possible? Is there no room for 2 molecules in the
> asymmetric unit, and the Patterson peak isa function of the "highly
> repetitive dimer"
> Eleanor
> If that is so you need to set the occupancy of any differences in the
> solution to 0.00 and check from the maps after refinement if you can see
> which copy of the molecule fits the difference density best - it would
> nice if you had a TRP/ALA pair of residues or something very distinctive..
> Eleanor


X Xiong, Cellular & Molecular Medicine wrote:
> Dear Crystallographers,
>
> We got a highly repetitive dimeric protein solved by SeMet-SAD in P21
> crystal form, and I am now trying to solve a dataset collected from a
> non-reproducible orthorhombic crystal of the same protein using the
> structure refined from P21 data.
>
>> From the Scala statistics, the orthorhombic crystal diffracted to
>> 2.2Å with
> an I/sigma of 3.1 at outmost shell; 98% complete overall, 89% complete
> 43.4-7.0Å, 99% complete 2.32-2.2Å, no twinning was detected. Due to
> the incompleteness at low resolution, it was hard to determine which
> orthorhombic space group it is in so data was scaled in P222. Very
> strong pseudo-translational symmetry has been detected by
> self-Patterson, as shown for reindexed data P21212 (space group later
> found by Phaser):
>
>  Order No. Site Height/Rms    Grid      Fractional coordinates
> Orthogonal coordinates
>
>     1    1    1  128.24     0   0   0   0.0000  0.0000  0.0000
> 0.00 0.00   0.00
>     2   13   13   57.51    60  44  38   0.5000  0.2741  0.5000
> 44.37 31.88  27.55
>     3    2    2   33.75     0   7   0   0.0000  0.0414  0.0000
> 0.00 4.82   0.00
>     4   14   14   16.09    60  50  38   0.5000  0.3150  0.5000
> 44.37 36.63  27.55
>     5   12   12   15.75    60  37  38   0.5000  0.2324  0.5000
> 44.37 27.03  27.55
>     6    3    3   12.28     0  13   0   0.0000  0.0836  0.0000
> 0.00 9.72   0.00
>     7   15    0    7.06    60  57  38   0.5000  0.3574  0.5000
> 44.37 41.56  27.55
>     8    4    4    6.18     0  72   0   0.0000  0.4503  0.0000
> 0.00 52.36   0.00
>     9    9    9    5.68     5   0   5   0.0410  0.0000  0.0683
> 3.64 0.00   3.76
>    10    5    5    5.36     2  20   2   0.0142  0.1254  0.0206
> 1.26 14.59   1.14
>    11   11   11    5.33    58  31  38   0.4852  0.1909  0.5000
> 43.06 22.20  27.55
>    12    6    6    3.98     5   0   2   0.0435  0.0000  0.0286
> 3.86 0.00   1.58
>    13    7    7    3.82     2  27   3   0.0168  0.1659  0.0334
> 1.49 19.30   1.84
>    14    8    8    3.68     0   0   5   0.0000  0.0000  0.0722
> 0.00 0.00   3.98
>    15   10   10    3.41    60  64  37   0.5000  0.4007  0.4872
> 44.37 46.59  26.84
>
> Phaser was used to test all possible alternative space groups to find
> MR solution using the structure from P21 data:
>
> #   Phaser_P222_MosFLM_all_spacegroup
> SPACegroup HALL  P 2bc 2 #P 2 21 21
> SOLU SET  RFZ=9.1 TFZ=24.3 PAK=0 LLG=2545 LLG=3718
> SOLU 6DIM ENSE ensemble1 EULER  273.097    1.162   88.144 FRAC
> -0.03394 0.50659 -0.22125
> SOLU SET  RFZ=9.1 TFZ=25.0 PAK=0 LLG=2496 LLG=3622
> SOLU 6DIM ENSE ensemble1 EULER   91.491    0.850   89.812 FRAC
> 0.03435 -0.00618  0.00352
>
> and it found 2 solutions with very similar Z-scores and LLG gains, If
> I am right they are not crystallographic equivalent, and Phaser checks
> that as well.
>
> I reindexed the data to P21212 and Phaser found the same solutions:
>
> #   Phaser_Reindexed_P21212_2_solutions
> SPACegroup HALL  P 2 2ab #P 21 21 2
> SOLU SET  RFZ=10.0 TFZ=23.0 PAK=0 LLG=3266 LLG=3718
> SOLU 6DIM ENSE ensemble1 EULER   88.843   90.063    1.249 FRAC
> -0.00661 -0.22126 -0.46598
> SOLU SET  RFZ=9.9 TFZ=23.2 PAK=0 LLG=3178 LLG=3624
> SOLU 6DIM ENSE ensemble1 EULER  270.841   89.977  181.339 FRAC
> -0.50634 -0.00350  0.46533
>
> The difference between the two solutions seems to be that the second
> solution translated along the longer 21 axis by about ~32Å, I chose
> the first solution to re-build and refine, and final R/Rfree I got was
> 21.6%/26.5%. After that, I hope to solve the ambiguity of which MR
> solution is right by running Phaser again with the complete model
> (including H2O):
>
> #   Phaser_Reindexed_P21212_2_solutions
> SPACegroup HALL  P 2 2ab #P 21 21 2
> SOLU SET  RFZ=12.8 TFZ=28.4 PAK=0 LLG=6542 LLG=7649
> SOLU 6DIM ENSE ensemble1 EULER  180.156    0.000    0.000 FRAC
> -0.50060 -0.00065  0.49998
> SOLU SET  RFZ=12.8 TFZ=32.3 PAK=0 LLG=6036 LLG=7059
> SOLU 6DIM ENSE ensemble1 EULER  179.201  180.000    0.000 FRAC
> 0.00227 0.22262 -0.50054
>
> It seems that the previous first solution has become the second
> solution, while the previous second solution became the first. Refmac
> refinement was performed on both solutions (H2O removed) came out from
> Phaser,
>
> solution 1 R/Rfree = 24.1/28.9
> solution 2 R/Rfree = 24.8/28.7
>
> the previous first solution got slightly worse scores, however, the
> R-factors for both solutions are so similar and both of them gave very
> similar electron density that I can not figure out which one is the
> right solution.
>
> I would very grateful for any advices.
>
> Thanks in advance,



----------------------
Xiaoli Xiong
PhD Candidate
Department of Cellular and Molecular Medicine
School of Medical Sciences
University of Bristol
University Walk
Bristol BS8 1TD, UK
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