[phenixbb] help on MIR dataset

Terwilliger, Thomas Charles terwilliger at lanl.gov
Fri Oct 24 15:05:20 PDT 2014


Hi Fengyun,

Oh right...I forgot you can only use the density from a map as phase information for SAD phasing.  Instead try these keywords:

input_phase_file=overall_best_denmod_map_coeffs_from6.mtz
input_phase_labels="FWT,PHWT"

Let me know if that doesn't do it!

All the best,
Tom T

________________________________________
From: Ni, Fengyun [fni at bcm.edu]
Sent: Friday, October 24, 2014 2:51 PM
To: Terwilliger, Thomas Charles
Cc: PHENIX user mailing list
Subject: RE: [phenixbb] help on MIR dataset

Hi Tom,

Thank you for your suggestions!

I did try your #1 advice with the old version phenix1.8. The resulting model does not make much sense.
I just install the official phenix 1.9 to run you #2 advice with the following commands but getting error message at the end of file. Please help me. Thanks again!

autosol {
  seq_file = ../seq-prot.dat
  crystal_info {
    solvent_fraction = 0.62
  }
  native {
    data = ../nat.mtz
  }
  deriv {
    data = ../pb1-all-4.4.mtz
    atom_type = Pb
    lambda = 0.9464
  }
  phasing {
    input_part_map_coeffs_file = overall_best_denmod_map_coeffs_from6.mtz
    input_part_map_coeffs_labels = "FWT,PHWT"
    phaser_sites_then_phase = True
  }
}


......
Set try_orig_sad_data_in_hyss=False as thoroughness=quick and extreme_dm=False
Setting ha_iteration to True by default as this is mir/mad
Not truncating ha sites at start of resolve as this is not PHASER SAD phasing

Sorry, you can only use an MR model to find sites with a single dataset and Phaser SAD phasing

Sorry: Sorry, you can only use an MR model to find sites with a single dataset and Phaser SAD phasing
------end of error message




________________________________________
From: Terwilliger, Thomas Charles [terwilliger at lanl.gov]
Sent: Friday, October 24, 2014 12:59 PM
To: Ni, Fengyun
Cc: PHENIX user mailing list; Terwilliger, Thomas Charles
Subject: RE: [phenixbb] help on MIR dataset

Hi Fengyun,

It sounds as though your Hg dataset is very promising.  Here's what I would try:

1. take the results from your Hg autosol run (overall_best_denmod_map_coeffs.mtz; overall_best_ha.pdb; overall_best_refine_data.mtz) and put them into autobuild along with hires_file=my_native_3A_data.mtz and your sequence file. (Perhaps you already tried this one).

2. Alternative: find the sites in the Pb using the Hg solution:   Run autosol with the Pb data and the keywords:
   input_part_map_coeffs_file=AutoSol_run_xxxx_/overall_best_denmod_map_coeffs.mtz  (xxx=your autosol run with Hg)
   phaser_sites_then_phase=True   (find sites using that map file, then phase only using the Pb phases)

  Then compare the map from the Hg with the Pb map with:
  phenix.get_cc_mtz_mtz AutoSol_run_xxxx_/overall_best_denmod_map_coeffs.mtz  AutoSol_run_yyyy_/overall_best_denmod_map_coeffs.mtz

  If they are similar (cc>0.2 or so) then you are on the right track with both maps.  You can combine them by running autosol one more time (with a parameters file something like this):

autosol {
  seq_file = seq.dat

  wavelength {
    group = 1
    data = Hg.sca
    lambda = 1.1
    atom_type = Hg
    f_prime = -7.
    f_double_prime = 4.5
    sites_file = hg.pdb
  }
  wavelength {
    group = 2
    data = Pb.sca
    lambda = 1.1
    atom_type = Pb
    f_prime = -7.
    f_double_prime = 4.5
    sites_file = pb.pdb
  }
}

All the best,
Tom T


________________________________________
From: Ni, Fengyun [fni at bcm.edu]
Sent: Friday, October 24, 2014 9:51 AM
To: Terwilliger, Thomas Charles
Cc: PHENIX user mailing list
Subject: RE: [phenixbb] help on MIR dataset

Hi Tom,

Thank you very much for the suggestions! It takes some time for me to try all your suggestions.

Now I could get a solution for a Hg-derivative dataset at very low resolution (5 A) with the following statistics,

                               SITE  ATOM  OCCUP     X           Y            Z             B
 CURRENT VALUES:      1    Hg       0.8672  0.0160  0.7559  0.2200   48.4721
 CURRENT VALUES:      2    Hg       1.0700  0.4392  0.3669  0.2884   60.0000
 CURRENT VALUES:      3    Hg       0.0196  0.5968  0.9263  0.1392    1.0000
 CURRENT VALUES:      4    Hg       0.1662  0.1408  0.6124  0.2366    6.4178
 CURRENT VALUES:      5    Hg       0.3109  0.1818  0.3807  0.2923   56.7720

Solution # 1   BAYES-CC: 52.8 +/- 20.8 (2SD) Dataset #1   FOM: 0.62
Score type:       SKEW    CORR_RMS    NCS_OVERLAP
Raw scores:        0.23      0.85      0.00
100x EST OF CC:   52.84     49.93     31.25

The resulting map looks reasonable for the fact that several long consecutive densities could be located and it somehow looks like what my protein should be like. My native data only diffract to 3.1 A, and the model building based on the above solution (after density modification) could only build some fragment without alpha/beta secondary structures, though R/Rfree is around 0.38/0.47.

I have another Pb-derivative, it could find a following solution at 4.4 A,

                            SITE  ATOM       OCCUP     X          Y       Z         B
 CURRENT VALUES:      1    Pb       0.2317  0.0643  0.6622  0.1180   60.0000
 CURRENT VALUES:      2    Pb       0.0534  0.9454  0.5850  0.2217    1.0000

Solution # 2  BAYES-CC: 54.3 +/- 19.2 (2SD) Dataset #1   FOM: 0.39
Score type:       SKEW    CORR_RMS    NCS_OVERLAP
Raw scores:        0.24      0.69      0.00
100x EST OF CC:   54.29     18.27     31.25

The map from Pb-derivative is not as good as that from Hg-derivative. And it could not be combined with Hg-solution when running autosol together for both datasets.

I read from phenix that there's autobuild_parallel version for building from bad maps with some options like building with BUCCANEER, does anyone have suggestions on running this? Any suggestions for what I could try next are welcome.

Thank you very much!
Fengyun


________________________________________
From: Terwilliger, Thomas Charles [terwilliger at lanl.gov]
Sent: Tuesday, August 26, 2014 10:13 AM
To: Ni, Fengyun
Cc: PHENIX user mailing list; Terwilliger, Thomas Charles
Subject: Re: [phenixbb] help on MIR dataset

Hi Fengyun,

Some things to try:

1. Are your data twinned (look at intensity moments in xtriage output)?

2. Try space groups P3121 (and P3221 and P321) in addition to P3.

3. Take your best MR solution (or any MR solutions) and calculate phases. Use those phases as input phases to autosol and it will try to find sites using those phases.  If you then get back a map that has correlation with these phases (use get_cc_mtz_mtz to check) then you probably have a solution (autosol will only use those input phases to find the sites, not in phasing).

4. Try each of your datasets separately as SAD datasets.  You can use MR SAD in autosol (similar to #2 above but using phaser to calculate phases).

All the best,
Tom T


On Aug 25, 2014, at 10:07 PM, Ni, Fengyun wrote:

> Hi everyone,
>
> I am working on a MIR dataset: one native data, three EMTS soaked data at different concentration, one PbAc2 data. My initial trial with phenix autosol did not give positive solutions. I hope I could get some information from all of you.
>
> I index and integrate the data in Mosflm and possible group is P3, though the Pointless said the most possible is P3121. But for now, I only tried P3 at different resolutions for the reason that a potential MR solution (below 20% identities, TFZ=7.1) was found in P31 space group. If P3121 is the real space group, the space is too crowed to fit my molecules unless they have some unexpected symmetry.
>
> For the difference among dataset in P3 space group, the output from SCALEIT of CCP4 gives as follows,
>
> RMS differences|RMSiso      RMSano
> ----------------------------------------
> FP-hg          |172.30      22.50
> FP-hg2         |159.20      24.07
> FP-hg3         |85.08       27.92
> FP-pb          |31.15       24.55
>
> The native data's resolution cutoff is 3.2 A, and 3.2/3.8/3.5 for different EMTS data, 3.5 for Pb data.I did notice some radiation damage for EMTS (Hg) data (large negative B-factors in SCALA), but not for Pb data. And the anomalous signal only extend to 5.6 and 4.5 A for EMTS and Pb data, respectively, as indicated by phenix.xtriage. These are all the possible heavy atom data we could obtain. So i tried all of them in phenix.autosol. The input is like follows,
>
> autosol {
>  seq_file = ../seq.dat
>  crystal_info {
>  solvent_fraction = 0.62  #determined from matthews coefficient to assure "reasonable" content in asymmetric unit.
>  }
>  native {
>    data = nat.mtz
>  }
>  deriv {
>    data = hg1.mtz
>    atom_type = Hg
>    inano = noinano *inano anoonly
>    lambda = 1.00394
>  }
>  deriv {
>    data = hg2.mtz
>    atom_type = Hg
>    inano = noinano *inano anoonly
>    lambda = 1.00394
>  }
>  deriv {
>    data = hg3.mtz
>    atom_type = Hg
>    inano = noinano *inano anoonly
>    lambda = 1.00394
>  }
>  deriv {
>    data = pb.mtz
>    atom_type = Pb
>    inano = noinano *inano anoonly
>    lambda = 0.94640
>  }
>  model_building {
>    build = False   #my structure is DNA/protein complex, so i choose to stop the building process.
> }
> }
>
> I tried at different resolutions, 3.6/4.0/4.4/4.8/5.2/5.6/6.0 A. One solution was given based on Pb data at 4.8 A with following statistics,
>
> Solution # 6  BAYES-CC: 59.7 +/- 17.2 (2SD) Dataset #4   FOM: 0.24
> Score type:       SKEW    CORR_RMS    NCS_OVERLAP
> Raw scores:        0.31      0.69      0.00
> 100x EST OF CC:   59.74     17.55     31.25
>
> Refined heavy atom sites (fractional):
>           X         Y          Z
> xyz       0.033      0.020      0.538
> xyz       0.417      0.217      0.113
> xyz       0.041      0.010      0.721
>
>
> The statistics is almost the same for its inverse, and i read from the output that the statistics for phasing other Hg datasets are very low (BAYES-CC is lower than 20) based on the above solution. The resulting map does not make much sense yet.
>
> Sorry for the long post. Any suggestion is welcome.
> Thank you very much!
> Fengyun
> _______________________________________________
> phenixbb mailing list
> phenixbb at phenix-online.org
> http://phenix-online.org/mailman/listinfo/phenixbb



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