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This is an example of S-SAD structure solution (PDB id [http://www.rcsb.org/pdb/explore.do?structureId=2QVO | This is an example of S-SAD structure solution (PDB id [http://www.rcsb.org/pdb/explore.do?structureId=2QVO 2QVO]), a 95-residue protein used by James Tucker Swindell II to establish optimized procedures for data reduction. The data available to solve the structure are two runs of 360° collected at a wavelength of 1.9Å. | ||
==XDS data reduction== | ==XDS data reduction== | ||
In the course of writing this up, it turned out that it was not necessary to scale the two datasets together, using [[XSCALE]], because the structure can be solved from any of the two, separately. | In the course of writing this up, it turned out that it was not necessary to scale the two datasets together, using [[XSCALE]], because the structure can be solved from any of the two, separately. But, of course, structure solution would be easier when merging the data (try for yourself!). | ||
===dataset 1=== | ===dataset 1=== | ||
Using | Using [[generate_XDS.INP]] "../../APS/22-ID/2qvo/ACA10_AF1382_1.0???" we obtain: | ||
<pre> | <pre> | ||
JOB= XYCORR INIT COLSPOT IDXREF DEFPIX INTEGRATE CORRECT | JOB= XYCORR INIT COLSPOT IDXREF DEFPIX INTEGRATE CORRECT | ||
| Line 73: | Line 73: | ||
* 21 tP 7.3 53.5 53.5 41.2 90.1 90.1 90.3 0 1 0 0 0 0 -1 0 -1 0 0 0 | * 21 tP 7.3 53.5 53.5 41.2 90.1 90.1 90.3 0 1 0 0 0 0 -1 0 -1 0 0 0 | ||
39 mC 249.8 114.5 41.2 53.5 90.1 90.3 69.0 1 -2 0 0 1 0 0 0 0 0 1 0 | 39 mC 249.8 114.5 41.2 53.5 90.1 90.3 69.0 1 -2 0 0 1 0 0 0 0 0 1 0 | ||
indicating at most tetragonal symmetry, | indicating at most tetragonal symmetry. Below this table, CORRECT calculates R-factors for each of the lattices whose metric symmetry is compatible with the cell of the crystal (marked by * in the table above): | ||
SPACE-GROUP UNIT CELL CONSTANTS UNIQUE Rmeas COMPARED LATTICE- | SPACE-GROUP UNIT CELL CONSTANTS UNIQUE Rmeas COMPARED LATTICE- | ||
NUMBER a b c alpha beta gamma CHARACTER | NUMBER a b c alpha beta gamma CHARACTER | ||
| Line 140: | Line 140: | ||
NUMBER OF UNIQUE ACCEPTED REFLECTIONS 13784 | NUMBER OF UNIQUE ACCEPTED REFLECTIONS 13784 | ||
So the anomalous signal goes to about 3.3 | So the anomalous signal goes to about 3.3 Å (which is where 30% would be, in the "Anomal Corr" column), and the useful resolution goes to 2.16 Å, I'd say (pls note that this table treats Friedels separately; merging them increases I/sigma by another factor of 1.41). | ||
For the sake of comparability, from now on we use the same axes (53.03 53.03 40.97) as the deposited PDB id 2QVO. | For the sake of comparability, from now on we use the same axes (53.03 53.03 40.97) as the deposited PDB id 2QVO. | ||
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==SHELXC/D/E structure solution== | ==SHELXC/D/E structure solution== | ||
This is done in a subdirectory of the XDS data reduction directory (of dataset "1" or "2"). Here, we use a script to generate XDSCONV.INP (I used MERGE=TRUE, sometimes the results are better that way), run [[XDSCONV|xdsconv]] and [[ccp4com:SHELX_C/D/E|SHELXC]]. | This is done in a subdirectory of the XDS data reduction directory (of dataset "1" or "2"). Here, we use a script to generate XDSCONV.INP (I used MERGE=TRUE, sometimes the results are better that way; update Sep 2011: the [[ccp4com:SHELX_C/D/E#Obtaining_the_SHELX_programs|beta-test version of SHELXC]] fixes this problem, so MERGE=FALSE would be preferable since it gives more statistics output), run [[XDSCONV|xdsconv]] and [[ccp4com:SHELX_C/D/E|SHELXC]]. | ||
<pre> | <pre> | ||
#!/bin/csh -f | #!/bin/csh -f | ||
| Line 266: | Line 266: | ||
shelxd j_fa | shelxd j_fa | ||
This gives best CC All/Weak of 37.28 / 21.38 for dataset 1, and best CC All/Weak of 37.89 / 23.80 for dataset 2 | The "FIND 3" needs a comment: the sequence has 4 Met and 1 Cys, but we don't expect to find the N-terminal Met. Since SHELXD always searches for more atoms than specified, we might as well tell it to try and locate 3 sulfurs. | ||
This gives best CC All/Weak of 37.28 / 21.38 for dataset 1, and best CC All/Weak of 37.89 / 23.80 for dataset 2. | |||
Next we run G. Sheldrick's beta-Version of [[ccp4com:SHELX_C/D/E|SHELXE]] Version 2011/1: | Next we run G. Sheldrick's beta-Version of [[ccp4com:SHELX_C/D/E|SHELXE]] Version 2011/1: | ||
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so the density is better, but not much. Furthermore, we note in passing that the number of anomalous scatterers (5) matches the sum of 4 Met and 1 Cys in the sequence. | so the density is better, but not much. Furthermore, we note in passing that the number of anomalous scatterers (5) matches the sum of 4 Met and 1 Cys in the sequence. | ||
== | ==Exploring the limits== | ||
With dataset 2, I tried to use the first 270 frames and could indeed solve the structure using the above SHELXC/D/E approach (with WFAC1=1.5) - 85 residues in a single chain, with "CC for partial structure against native data = 47.51 %". It should be mentioned that I also tried this in November 2009, and it didn't work with the version of XDS available then! | With dataset 2, I tried to use the first 270 frames and could indeed solve the structure using the above SHELXC/D/E approach (with WFAC1=1.5) - 85 residues in a single chain, with "CC for partial structure against native data = 47.51 %". It should be mentioned that I also tried this in November 2009, and it didn't work with the version of XDS available then! | ||
With 180 frames, it was possible to get a complete model by (twice) re-cycling the j.hat file to j_fa.res. '''This means that the structure can be automatically solved just from the first 180 frames of dataset 2!''' | With 180 frames, it was possible to get a complete model by (twice) re-cycling the j.hat file to j_fa.res. '''This means that the structure can be automatically solved just from the first 180 frames of dataset 2!''' | ||
==Availability== | |||
* [https://{{SERVERNAME}}/pub/xds-datared/2qvo/xds-2qvo-1-1_360-F.mtz] - amplitudes for frames 1-360 of dataset 1. | |||
* [https://{{SERVERNAME}}/pub/xds-datared/2qvo/xds-2qvo-1-1_360-I.mtz] - intensities for frames 1-360 of dataset 1. | |||
* [https://{{SERVERNAME}}/pub/xds-datared/2qvo/xds-2qvo-2-1_180-F.mtz] - amplitudes for frames 1-180 of dataset 2. | |||
* [https://{{SERVERNAME}}/pub/xds-datared/2qvo/xds-2qvo-2-1_180-I.mtz] - intensities for frames 1-180 of dataset 2. | |||
* [https://{{SERVERNAME}}/pub/xds-datared/2qvo/xds-2qvo-2-1_360-F.mtz] - amplitudes for frames 1-360 of dataset 2. | |||
* [https://{{SERVERNAME}}/pub/xds-datared/2qvo/xds-2qvo-2-1_360-I.mtz] - intensities for frames 1-360 of dataset 2. | |||
As you can see, all these files are in the same directory [https://{{SERVERNAME}}/pub/xds-datared/2qvo/]. I put there the XDS_ASCII.HKL files and SHELXD/SHELXE result files as well. | |||