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centric reflections

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  • Etienne Ladoine
    Dear all, I recently collected SAD data on a SeMet subsituted protein. Resolution is 1.6 Angstrom, data 99% complete, spacegroup P212121. 8 selenium sites
    Message 1 of 2 , Feb 4, 2002
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      Dear all,

      I recently collected SAD data on a SeMet subsituted protein.
      Resolution is 1.6 Angstrom, data 99% complete, spacegroup P212121.
      8 selenium sites could readily be found using CNS. However, after phasing I
      get the statistics shown below. The Rcullis has a ridiculous (impossible?)
      value for the centric reflections, and the FOM is very low for the centrics
      as well, compared to the overal FOM.
      What can be the reason for this huge difference between centrics and
      acentrics ?
      After density modification and choosing the proper hand, the maps look quite
      OK, with the main chain readily traceable by Arp-wArp...

      Etienne Ladoine


      the statistics:


      generalized Cullis R-value
      --------------------------
      sqrt<loc^2>/sqrt<delta F^2>
      centrics acentrics all
      phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
      #bin | resolution range | #refl |
      1 3.18 27.92 13742 10.0000 0.4493 0.4708
      2 2.52 3.18 14011 9.3071 0.4442 0.4578
      3 2.20 2.52 13807 7.4640 0.4573 0.4692
      4 2.00 2.20 13666 6.5925 0.4699 0.4808
      5 1.86 2.00 13512 6.9384 0.4780 0.4882
      6 1.75 1.86 13248 6.9793 0.4863 0.4961
      7 1.66 1.75 12996 6.8612 0.5038 0.5136
      8 1.59 1.66 10892 6.9733 0.5153 0.5247
      #bin | resolution range | #refl |
      1 1.59 27.92 105874 8.8088 0.4649 0.4787


      Kraut R-value
      -------------
      sum|(|Fp+Fh|-|Fph|)|/sum|Fp|
      centrics acentrics all
      phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
      #bin | resolution range | #refl |
      1 3.18 27.92 13742 0.1019 0.0470 0.0514
      2 2.52 3.18 14011 0.1068 0.0545 0.0573
      3 2.20 2.52 13807 0.1229 0.0599 0.0627
      4 2.00 2.20 13666 0.1260 0.0614 0.0641
      5 1.86 2.00 13512 0.1429 0.0712 0.0739
      6 1.75 1.86 13248 0.1487 0.0809 0.0834
      7 1.66 1.75 12996 0.1662 0.0880 0.0908
      8 1.59 1.66 10892 0.1840 0.0944 0.0973
      #bin | resolution range | #refl |
      1 1.59 27.92 105874 0.1187 0.0623 0.0653
      -----------------------------------------------------------------


      phasing power
      -------------
      sqrt<Fh^2>/sqrt<var_loc^2>
      centrics acentrics all
      phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
      #bin | resolution range | #refl |
      1 3.18 27.92 13742 2.8250 2.7798 2.7839
      2 2.52 3.18 14011 2.6190 2.7606 2.7526
      3 2.20 2.52 13807 2.8262 2.7514 2.7552
      4 2.00 2.20 13666 2.7100 2.5668 2.5734
      5 1.86 2.00 13512 2.5707 2.5162 2.5185
      6 1.75 1.86 13248 2.3787 2.4793 2.4754
      7 1.66 1.75 12996 2.4720 2.4236 2.4254
      8 1.59 1.66 10892 2.5127 2.4391 2.4418
      #bin | resolution range | #refl |
      1 1.59 27.92 105874 2.7060 2.6484 2.6517
      ---------------------------------------------------------------



      figure-of-merit
      ---------------
      centrics acentrics all
      phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
      #bin | resolution range | #refl |
      1 3.18 27.92 13742 0.1296 0.5210 0.4859
      2 2.52 3.18 14011 0.1390 0.5218 0.4994
      3 2.20 2.52 13807 0.1549 0.5135 0.4954
      4 2.00 2.20 13666 0.1528 0.5085 0.4924
      5 1.86 2.00 13512 0.1584 0.5043 0.4899
      6 1.75 1.86 13248 0.1478 0.4953 0.4817
      7 1.66 1.75 12996 0.1672 0.4852 0.4731
      8 1.59 1.66 10892 0.1563 0.4728 0.4615
      #bin | resolution range | #refl |
      1 1.59 27.92 105874 0.1473 0.5036 0.4857
      ---------------------------------------------------------------






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    • Poul Nissen
      Centric reflections contain no anomalous signal - only your acentric reflections have been phased by SAD. You should just forget about those weired statistics
      Message 2 of 2 , Feb 4, 2002
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        Centric reflections contain no anomalous signal - only your acentric
        reflections have been phased by SAD.
        You should just forget about those weired statistics on the centric reflections
        - they have no meaning.
        DM will nail the phases of the centric reflections, if a decent envelope can be
        established beginning from the SAD phases. It certainly seems that your SAD
        experiment was very succeful!

        For a good example of how to get the very most out of your SAD experiment, and
        thus the best possible experimental map, see:
        Vestergaard B, Van LB, Andersen GR, Nyborg J, Buckingham RH, Kjeldgaard M.
        Bacterial polypeptide release factor RF2 is structurally distinct from
        eukaryotic eRF1.
        Mol Cell. 2001 Dec;8(6):1375-82.



        Poul


        Etienne Ladoine wrote:

        > Dear all,
        >
        > I recently collected SAD data on a SeMet subsituted protein.
        > Resolution is 1.6 Angstrom, data 99% complete, spacegroup P212121.
        > 8 selenium sites could readily be found using CNS. However, after phasing I
        > get the statistics shown below. The Rcullis has a ridiculous (impossible?)
        > value for the centric reflections, and the FOM is very low for the centrics
        > as well, compared to the overal FOM.
        > What can be the reason for this huge difference between centrics and
        > acentrics ?
        > After density modification and choosing the proper hand, the maps look quite
        > OK, with the main chain readily traceable by Arp-wArp...
        >
        > Etienne Ladoine
        >
        > the statistics:
        >
        > generalized Cullis R-value
        > --------------------------
        > sqrt<loc^2>/sqrt<delta F^2>
        > centrics acentrics all
        > phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
        > #bin | resolution range | #refl |
        > 1 3.18 27.92 13742 10.0000 0.4493 0.4708
        > 2 2.52 3.18 14011 9.3071 0.4442 0.4578
        > 3 2.20 2.52 13807 7.4640 0.4573 0.4692
        > 4 2.00 2.20 13666 6.5925 0.4699 0.4808
        > 5 1.86 2.00 13512 6.9384 0.4780 0.4882
        > 6 1.75 1.86 13248 6.9793 0.4863 0.4961
        > 7 1.66 1.75 12996 6.8612 0.5038 0.5136
        > 8 1.59 1.66 10892 6.9733 0.5153 0.5247
        > #bin | resolution range | #refl |
        > 1 1.59 27.92 105874 8.8088 0.4649 0.4787
        >
        > Kraut R-value
        > -------------
        > sum|(|Fp+Fh|-|Fph|)|/sum|Fp|
        > centrics acentrics all
        > phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
        > #bin | resolution range | #refl |
        > 1 3.18 27.92 13742 0.1019 0.0470 0.0514
        > 2 2.52 3.18 14011 0.1068 0.0545 0.0573
        > 3 2.20 2.52 13807 0.1229 0.0599 0.0627
        > 4 2.00 2.20 13666 0.1260 0.0614 0.0641
        > 5 1.86 2.00 13512 0.1429 0.0712 0.0739
        > 6 1.75 1.86 13248 0.1487 0.0809 0.0834
        > 7 1.66 1.75 12996 0.1662 0.0880 0.0908
        > 8 1.59 1.66 10892 0.1840 0.0944 0.0973
        > #bin | resolution range | #refl |
        > 1 1.59 27.92 105874 0.1187 0.0623 0.0653
        > -----------------------------------------------------------------
        >
        > phasing power
        > -------------
        > sqrt<Fh^2>/sqrt<var_loc^2>
        > centrics acentrics all
        > phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
        > #bin | resolution range | #refl |
        > 1 3.18 27.92 13742 2.8250 2.7798 2.7839
        > 2 2.52 3.18 14011 2.6190 2.7606 2.7526
        > 3 2.20 2.52 13807 2.8262 2.7514 2.7552
        > 4 2.00 2.20 13666 2.7100 2.5668 2.5734
        > 5 1.86 2.00 13512 2.5707 2.5162 2.5185
        > 6 1.75 1.86 13248 2.3787 2.4793 2.4754
        > 7 1.66 1.75 12996 2.4720 2.4236 2.4254
        > 8 1.59 1.66 10892 2.5127 2.4391 2.4418
        > #bin | resolution range | #refl |
        > 1 1.59 27.92 105874 2.7060 2.6484 2.6517
        > ---------------------------------------------------------------
        >
        > figure-of-merit
        > ---------------
        > centrics acentrics all
        > phasing statistics of FOBS_FRIED (LOC phase set, excluding outliers)
        > #bin | resolution range | #refl |
        > 1 3.18 27.92 13742 0.1296 0.5210 0.4859
        > 2 2.52 3.18 14011 0.1390 0.5218 0.4994
        > 3 2.20 2.52 13807 0.1549 0.5135 0.4954
        > 4 2.00 2.20 13666 0.1528 0.5085 0.4924
        > 5 1.86 2.00 13512 0.1584 0.5043 0.4899
        > 6 1.75 1.86 13248 0.1478 0.4953 0.4817
        > 7 1.66 1.75 12996 0.1672 0.4852 0.4731
        > 8 1.59 1.66 10892 0.1563 0.4728 0.4615
        > #bin | resolution range | #refl |
        > 1 1.59 27.92 105874 0.1473 0.5036 0.4857
        > ---------------------------------------------------------------
        >
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        --
        ===================================================================
        Poul Nissen, Assoc. Research. Prof.
        Department of Molecular and Structural Biology
        Aarhus University nissen@...
        Gustav Wieds Vej 10c Tel. +45 8942 5024
        DK - 8000 Aarhus C, Denmark Fax +45 8612 3178
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