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(You are AnonymousGnome)

Content:

Introduction

See also:

If you want to install the software on your own computer(s) (Linux or MacOS): SHARP/autoSHARP home page - free for academics.

SHARP/autoSHARP tutorial (Friday, 3rd Dec 2021, 14:30-15:30)

We are going to follow one of the examples given here. In case you have trouble downloading the data files from our webpage, you should also be able to copy them from DLS computers:

  • open a Terminal in your NoMachine session (and make the window maybe a bit bigger, but not filling the full screen);
  • execute the following commands (you can cut-n-paste them):

      module load ccp4-workshop
      clusterme
      module load ccp4-workshop
 
      mkdir -p SHARP
      cd SHARP
      cp /dls/i04/data/2021/mx30951-2/processing/hcf00057/Workshop/SHARP/* .
      ls -l

1O22

A very quick example (from the larger list of examples) would be 1O22: 

  run_autoSHARP.sh \
    -fast -nowarp \
    -seq 1o22.pir -ha "Se" \
    -wvl 0.9778 peak -7 5 -sca 1o22_peak.sca \
    -d autoSHARP_SAD-1 | tee autoSHARP_SAD-1.lis

You should then see something like this:

run_autoSHARP_1.png

pointing you to the output file (a HTML file with the name LISTautoSHARP.html). Please open this file in a browser (e.g. Firefox) to see the full autoSHARP log file and results. You will some results regarding data analysis and substructure solution (with SHELXC/D):

run_autoSHARP_2a.png

and HA refinement and phasing (with SHARP), density modification (SOLOMON) and model-building (PARROT+BUCCANEER):

run_autoSHARP_2b.png

Of course, this is a nice simple SAD example that runs very fast:

run_autoSHARP_3.png

(it is nearly always the density-modification and building stage that takes the vast amount of running time).

At various times the HTML file will mention results being available - eg.

run_autoSHARP_4.png

You can cut-n-paste the text from the white box (including opening and closing brackets!) into a terminal window. At the end of the run, you will also see a summary on standard output (i.e. the Terminal where you executed the run_autoSHARP.sh command).

Insulin

If you want to use some of the Insulin datasets collected for this course:

  • First get the relevant sequence and one of those MTZ files, e.g. via
      cp /dls/i04/data/2021/mx30951-2/processing/hcf00057/Workshop/SHARP/ins.seq .
      cp /dls/i03/data/2021/mx30951-8/processed/gw/20211118/TestInsulin/ins_16/ins_16_2/autoPROC/staraniso_alldata-unique.mtz .
  • No need to run to the full resolution - solving a structure with 2A data should be fine (automatic building doesn't need higher resolution).
  • For a quick result let's add "–fast –nowarp".
  • So the total command could be
      module load ccp4-workshop
      clusterme
      module load ccp4-workshop
 
      mkdir -p SHARP
      cd SHARP

      run_autoSHARP.sh \
        -seq ins.seq \
        -ha S \
        -spgr I213 \
        -wvl 1.23987 peak \
        -mtz staraniso_alldata-unique.mtz \
        -R 50.0 2.0 -fast -nowarp \
        -d ins | tee ins.lis

Some tutorial-specific notes

Although we would always recommend to start with a run of autoSHARP using all defaults, for these workshop examples we are going to use a few extra flags: 

  run_autoSHARP.sh -nowarp ...

Although ARP/wARP building would potentially result in a much better built model (not necessary more complete), it doesn't scale with multiple processors/threads and therefore would overrun the tutorial time. You could also use 

  run_autoSHARP.sh -nowarp -fast ...

or even 

  run_autoSHARP.sh -nobuild -fast ...

if you are only interested in experimental phasing up to the first (hopefully interpretable) electron density map.

Where are my results?

At the end you should be presented (on standard output, i.e. within the Terminal) with something like 

 RESULTS:
 
   [01] HA detection (SHELXC/D) :
          initial sites = MAD_3ISY.01/SHELX/1.1.hatom
 
   [02] HA refinement and phasing (SHARP, first run) :
          refined sites = MAD_3ISY.01/SHARP.1/hatom.pdb
          phases        = MAD_3ISY.01/SHARP.1/eden.mtz
          residual maps = MAD_3ISY.01/SHARP.1/resid.mtz
 
   [03] HA refinement and phasing (SHARP, final run) :
          refined sites = MAD_3ISY.01/SHARP.3/hatom.pdb
          phases        = MAD_3ISY.01/SHARP.3/eden.mtz
          residual maps = MAD_3ISY.01/SHARP.3/resid.mtz

          eden.mtz:  see $SHARP_home/docs/sharp/manual/appendix1.html#eden
          resid.mtz: see $SHARP_home/docs/sharp/manual/appendix1.html#resid
 
   [04] First density modification (after hand-determination) :
          phases = MAD_3ISY.01/SHARP.3/eden_flat_45.1pc.mtz
 
          see $SHARP_home/docs/sharp/manual/appendix1.html#edenflat for details
          (use FBshasol/PHIBshasol for map calculation or viewing)
 
   [05] First model (built into initial density-modified map) :
          model  = MAD_3ISY.01/SHARP.3/LJS_45.1pc/buccaneer_cycle-5.pdb
          phases = MAD_3ISY.01/SHARP.3/LJS_45.1pc/parrot_cycle-5.mtz
 
          (use parrot.F_phi.F/parrot.F_phi.phi for map calculation or viewing)
 
   [06] Final density modification (after inclusion of previously built models) :
          phases = MAD_3ISY.01/SHARP.3/eden_flat_44.8pc.mtz
 
          see $SHARP_home/docs/sharp/manual/appendix1.html#edenflat for details
          (use FBshasol/PHIBshasol for map calculation or viewing)
 
   [07] Final model (built into last density-modified map) :
          model  = MAD_3ISY.01/SHARP.3/LJS_44.8pc/buccaneer_cycle-5.pdb
          phases = MAD_3ISY.01/SHARP.3/LJS_44.8pc/parrot_cycle-5.mtz
 
          (use parrot.F_phi.F/parrot.F_phi.phi for map calculation or viewing)
 
 NOTE: results are also collected in directory MAD_3ISY.01/Results
 
   -rw-rw----+ 1 hcf00057 mx27960_4 85 Dec  2 13:01 MAD_3ISY.01/Results/01_1.1.hatom
 
   -rw-rw----+ 1 hcf00057 mx27960_4   1014 Dec  2 13:18 MAD_3ISY.01/Results/02_coot.scm
   -rw-rw----+ 1 hcf00057 mx27960_4 509268 Dec  2 13:02 MAD_3ISY.01/Results/02_eden.mtz
   -rw-rw----+ 1 hcf00057 mx27960_4    366 Dec  2 13:02 MAD_3ISY.01/Results/02_hatom.pdb
   -rw-rw----+ 1 hcf00057 mx27960_4 355204 Dec  2 13:02 MAD_3ISY.01/Results/02_resid.mtz
 
   -rw-rw----+ 1 hcf00057 mx27960_4   1014 Dec  2 13:18 MAD_3ISY.01/Results/03_coot.scm
   -rw-rw----+ 1 hcf00057 mx27960_4 509268 Dec  2 13:02 MAD_3ISY.01/Results/03_eden.mtz
   -rw-rw----+ 1 hcf00057 mx27960_4    366 Dec  2 13:02 MAD_3ISY.01/Results/03_hatom.pdb
   -rw-rw----+ 1 hcf00057 mx27960_4 355128 Dec  2 13:02 MAD_3ISY.01/Results/03_resid.mtz
 
   -rw-rw----+ 1 hcf00057 mx27960_4    417 Dec  2 13:18 MAD_3ISY.01/Results/04_coot.scm
   -rw-rw----+ 1 hcf00057 mx27960_4 509668 Dec  2 13:08 MAD_3ISY.01/Results/04_eden_flat_45.1pc.mtz
   -rw-rw----+ 1 hcf00057 mx27960_4    366 Dec  2 13:02 MAD_3ISY.01/Results/04_hatom.pdb
 
   -rw-rw----+ 1 hcf00057 mx27960_4   78574 Dec  2 13:10 MAD_3ISY.01/Results/05_buccaneer_cycle-5.pdb
   -rw-rw----+ 1 hcf00057 mx27960_4     490 Dec  2 13:18 MAD_3ISY.01/Results/05_coot.scm
   -rw-rw----+ 1 hcf00057 mx27960_4 1112676 Dec  2 13:09 MAD_3ISY.01/Results/05_parrot_cycle-5.mtz
 
   -rw-rw----+ 1 hcf00057 mx27960_4    417 Dec  2 13:18 MAD_3ISY.01/Results/06_coot.scm
   -rw-rw----+ 1 hcf00057 mx27960_4 508968 Dec  2 13:16 MAD_3ISY.01/Results/06_eden_flat_44.8pc.mtz
 
   -rw-rw----+ 1 hcf00057 mx27960_4   79465 Dec  2 13:18 MAD_3ISY.01/Results/07_buccaneer_cycle-5.pdb
   -rw-rw----+ 1 hcf00057 mx27960_4     490 Dec  2 13:18 MAD_3ISY.01/Results/07_coot.scm
   -rw-rw----+ 1 hcf00057 mx27960_4 1111336 Dec  2 13:18 MAD_3ISY.01/Results/07_parrot_cycle-5.mtz
 

 You should be able to run Coot with the supplied scripts, e.g.:

   cd MAD_3ISY.01/Results
   coot --script 07_coot.scm

giving you very detailed information about the results (what they represent, how to view them and generally what to do with them). Further down you see a suggestion about refinement: 

 Files ready for refinement are:
   MAD_3ISY.01/Results/model.pdb
   MAD_3ISY.01/Results/for_refinement.mtz

 e.g. using BUSTER with

   refine -p MAD_3ISY.01/Results/model.pdb \
          -m MAD_3ISY.01/Results/for_refinement.mtz \
          -autoncs \
          -d BUSTER.01 | tee BUSTER.01.lis

 followed by

   coot --pdb BUSTER.01/refine.pdb --auto BUSTER.01/refine.mtz
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