Advanced Crystal Structure Analysis Using XDS with Small Molecule Data. Tim Grüne Georg-August-Universität Institut für Strukturchemie

Transcription

1 Advanced Crystal Structure Analysis Using XDS with Small Molecule Data Tim Grüne Georg-August-Universität Institut für Strukturchemie tt s 1 7t tg@shelx.uni-ac.gwdg.de November 13, 2013 Tim Grüne XDS: small molecule data 1/39

2 Overview XDS Availability and Documentation XDS Basics: P and the -card Parameter Refinement Indexing: Difficult cases like multiple lattices, poor diffraction 2 Small Molecule Examples: Synchrotron / data (s r t s) Tim Grüne XDS: small molecule data 2/39

3 The XDS Resources Author W. Kabsch Now distributed by K. Diederichs availability: XDS tt 1 s r main program suite Wiki and auxiliary programs tt str st 3 1 s GUIs xdsgui tt str st 3 1 s 1 very good graphical display of statistics Xdsapp tt t3 r rs s t tt r rs ss2 1 1 s optimised for data collection at BESSY - fast for standard cases. Tim Grüne XDS: small molecule data 3/39

4 Documentation Very well organised html-documentation Every STEP documented Every keyword documented Coordinate systems are explained Tim Grüne XDS: small molecule data 4/39

5 XDS Programs xds par Main program for data integration xscale par scaling program for multiple datasets cellparam Weighted average unit cell parameters from several runs Tim Grüne XDS: small molecule data 5/39

6 Templates Templates of input scripts for all supported detector formats Only very few adjustments necessary to get started Beamlines often generate appropriate input scripts r t P from XDSwiki for MARCCD, ADSC, and the Pilatus 6M It is worth learning how to set it up from scratch! tt str st 3 1 s 1 r t P Tim Grüne XDS: small molecule data 6/39

7 Benefits and Malefits Benefits robust indexing and robust integration integrate synchrotron data fast! (parallelised) optimised for Dectris Pilatus Detectors easy to fine-tune parameters Malefits not dedicated to twinned data or multiple lattices (treatment of overlaps) manual setup little graphical output no global parameter refinement for multiple runs Tim Grüne XDS: small molecule data 7/39

8 Program Control: P XDS is controlled by one single input file: P. Contains about 100 Keywords of the form Only about 10 Keywords must be modified for most data sets Most important one: P P P Each name stands for one of the steps XDS carries out during data integration. ( P is optional and corresponds to the BEST [2] or STRATEGY [1] output to report optimal data collection range(s)) Name cannot be changed Each data set must be run in separate directory to avoid overwriting of files. Tim Grüne XDS: small molecule data 8/39

9 Setting up P (starting from template) r s str P s 1 t t r r 1 s Direct beam position at 2θ = 0 Tim Grüne XDS: small molecule data 9/39

10 Steps of Data Processing Step (Important) input files output files XYCORR INIT COLSPOT IDXREF SPOT.XDS SPOT.XDS XPARM.XDS FRAME.cbf SPOT.XDS red: Text files with parameters or data cyan: control images (use 1 s r) black: data files for further processing DEFPIX BKGPIX.cbf ABS.cbf INTEGRATE XPARM.XDS INTEGRATE.HKL FRAME.cbf CORRECT INTEGRATE.HKL XDS_ASCII.HKL GXPARM.XDS Tim Grüne XDS: small molecule data 10/39

11 The Steps writes files for positional corrections of the detector plane. Most modern detectors provide already corrected images so that these to files are normally flat (0). determines initial detector background P collect strong spots for indexing ( P ) indexing: unit cell dimensions and crystal orientation P set active dectector area (exclude resolution cut-off, beam stop shadow, ice rings... ) P (optional) generate strategy tables with data completeness extract reflection intensities from frames applies corrections (polarisation, Lorentz-correction,... ), scales reflections, reports data statistics The ed is already scaled. Scaling of multiple data sets best done with. Tim Grüne XDS: small molecule data 11/39

12 Program Flow Each step must be passed at least once - the subsequent steps depend on files produced by the previous steps. Each step creates a log-file ( P P ). is the main hurdle - once unit cell and crystal orientation are determined, integration usually runs smoothly. summarises the quality of the data. Mostly P and P should be inspected. 1 r reads both the output after ( ) and the output from 1s r Tim Grüne XDS: small molecule data 12/39

13 : Multiple Runs Simple input script P: P P P First INPUT FILE: Reference and source for cell and spacegroup Usually no further options required Tim Grüne XDS: small molecule data 13/39

14 P Logfile P: statistcs table similar to 1 r P P P Tim Grüne XDS: small molecule data 14/39

15 P P P s P s r 1 t rr t t Tim Grüne XDS: small molecule data 15/39

16 Parameter Refinement Tim Grüne XDS: small molecule data 16/39

17 -ment of Parameters Refinement occurs at three stages: 1. : Initial parameter (Distance, beam, cell... ) 2. : Refinement of parameters per wedge of data ( P : macromolecular cell (default); P : small cell) 3. : Global refinement Default: all; reasonable setting: Tim Grüne XDS: small molecule data 17/39

18 Parameter Refinement Step (Important) input files output files XYCORR INIT globally refines the parameters ( based on all reflections COLSPOT IDXREF DEFPIX SPOT.XDS SPOT.XDS XPARM.XDS FRAME.cbf SPOT.XDS BKGPIX.cbf ABS.cbf P P 1 s r Do not run IDXREF: overwrite XPARM.XDS: P Automatic Laue group assignment unless P P present in P INTEGRATE XPARM.XDS INTEGRATE.HKL FRAME.cbf CORRECT INTEGRATE.HKL XDS_ASCII.HKL GXPARM.XDS Tim Grüne XDS: small molecule data 18/39

19 : Indexing Tim Grüne XDS: small molecule data 19/39

20 : Indexing 1. Input: strong spots found in P step 2. Saving time: Run COLSPOT on entire data set, select images for indexing with (multiple) P. Rerun, but not P when there are difficulties. Tim Grüne XDS: small molecule data 20/39

21 : Indexing Indexing step: Find cell parameters and cell orientation. First refinement of experimental parameters (Detector distance,... ) Writes solution to P P r st Tim Grüne XDS: small molecule data 21/39

22 P P : Detector coordinates and Intensity of strong spots to be used for indexing: 1 1 ts : Miller-Indices according to P 1 1 ts : not indexed with current cell Tim Grüne XDS: small molecule data 22/39

23 Failed Indexing XDS stops if less than 50 % of all reflections in SPOT.XDS could be indexed. P P P P P P Refined distance reasonable? Cell reasonable? P r Tim Grüne XDS: small molecule data 23/39

24 Example: Difficult Case P P P Tim Grüne XDS: small molecule data 24/39

25 Manual Indexing with 1 [3] P P : integrate with this solution Tim Grüne XDS: small molecule data 25/39

26 Twinning / Multiple Lattices: Search for Extra Lattices Patient? use s ts and Impatient? 1. create new directory r tt 2. r P tt P 3. rerun 4. compare orientations in P 5. integrate each lattice separately Caveat: is not set up to integrate overlapping spots Tim Grüne XDS: small molecule data 26/39

27 Example: Insulin Crystal with a Satellite a = (73.15, 24.82, 14.45) b = (10.46,59.86, 49.83) c = (26.74, 44.46, 59.02) a = (72.17, 27.40, 16.02) b = (30.84,51.13,51.48) c = ( 7.50, 53.39, 57.53) Tim Grüne XDS: small molecule data 27/39

28 Study Case I: SPAnH (R. Herbst-Irmer) Question: Are observed differences between F obs and F calc s t specific? 5 2θ = θ = 78 data converted from s r to r format Tim Grüne XDS: small molecule data 28/39

29 Setting up P for s r -format Difficulty: transfer collection geometry to XDS input description. t Coordinate System 5 5 r Coordinate System 5 5 r Non-perpendicular axes: rotation axis ( 100) or (0 10)... SPAnH: only ω scans: rotation axis = (0 10) Tim Grüne XDS: small molecule data 29/39

30 s r t s s r t s s r t t ss rt s 1 s str t t t 1 s Append to XDS.INP does not read sfrm (missing spatial correction) export to r within P Tim Grüne XDS: small molecule data 30/39

31 Unstable Refinement Small Cell = few spots within 5 Leads to unstable parameter refinement in Solution then: Increase P (20-60) Better: Ensure correct orientation after indexing and set no refinement during integration, global refinement of all parameters with all reflections Caveat: Check XDS can loose orientation and spots Tim Grüne XDS: small molecule data 31/39

32 Some Statistics for SPAnH s t t r2 t 2 s r s r st r s t str s r s P r s r t r t rs r s r str ts Tim Grüne XDS: small molecule data 32/39

33 Study Case II: Peroxo (K. Dalle, AK Meyer) Cell: Disorder Multiple lattices Five crystals +κ Goniometer at BL14.1 (BESSY): 10 Data sets collected, 199GB data, frames Tim Grüne XDS: small molecule data 33/39

34 XDS: Mosaicity and Beam Divergence Two Parameters control the size of spots on the detector: r t s t t r t r r ss t s r s rt st r t t s t2 If undefined: adjusted by during step Profiles shown in P search for " " Tim Grüne XDS: small molecule data 34/39

35 Undefined Mosaicity and Beam Divergence Combination of Spots Wrong Intensities large R-values Wrong Spot Positions: Wrong Parameter Refinement Jump between two lattices Tim Grüne XDS: small molecule data 35/39

36 Mosaicity and Beam Divergence too Small Wrong Intensities large R-values Inbetween two lattices: zero intensity Tim Grüne XDS: small molecule data 36/39

37 Mosaicity and Beam Divergence adjusted Tim Grüne XDS: small molecule data 37/39

38 One crystal, four κ settings: 0,10,20,30 Peroxo: Some Results P r s r t r t rs r s r str ts Tim Grüne XDS: small molecule data 38/39

39 References 1. Leslie, A.G.W., Recent changes to the MOSFLM package for processing film and image plate data, Joint CCP4 + ESF-EAMCB Newsletter on Protein Crystallography (1992), No A.N. Popov and Bourenkov, Choice of data-collection parameters based on statistic modeling Acta Crystallogr. (2003). D59, Andrew Arvai, arvai/adxv.html Tim Grüne XDS: small molecule data 39/39