Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results...
Transcript of Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results...
![Page 1: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/1.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data CollectionA New Synchrotron Serial Crystallography Method
Ulrich Zander
ESRF (Grenoble, France)
Users’Meeting 2015
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Introduction Method Test Cases First Results Perspectives
Acknowledgements
� EMBL Hamburg
Gleb Bourenkov
� ESRF
Alexander PopovChristoph Muller-DieckmannOlof SvensonDaniele De SanctisGordon Leonard
� IBS
Ivan ChushchinValentin GordeliyEkatarina Round
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Introduction Method Test Cases First Results Perspectives
Introduction
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Introduction Method Test Cases First Results Perspectives
Serial Crystallography
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Introduction Method Test Cases First Results Perspectives
Serial Crystallography
Serial collection of partial datasets of different crystal(position)s
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Introduction Method Test Cases First Results Perspectives
Serial Crystallography
Serial collection of partial datasets of different crystal(position)s
Kendrew et al. (1960) Structure of Myoglobin
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Introduction Method Test Cases First Results Perspectives
SFX: Serial femtosecond crystallography
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Introduction Method Test Cases First Results Perspectives
SFX: Serial femtosecond crystallography
Femtosecond nanocrystallography (Chapman et al., Nature 2011)
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Introduction Method Test Cases First Results Perspectives
SSX: Synchrotron serial crystallography
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Introduction Method Test Cases First Results Perspectives
SSX: Synchrotron serial crystallography
Structure of Trypanosoma brucei Procathepsin B at 3A
Gati et al., IUCrJ 2014: Serial crystallography on in vivo grownmicrocrystals using synchrotron radiation
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Introduction Method Test Cases First Results Perspectives
SSX: Synchrotron serial crystallography
Structure of Trypanosoma brucei Procathepsin B at 3A
Gati et al., IUCrJ 2014: Serial crystallography on in vivo grownmicrocrystals using synchrotron radiation
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Introduction Method Test Cases First Results Perspectives
Method
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Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
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Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 15: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/15.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 16: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/16.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 17: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/17.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 18: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/18.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 19: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/19.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 20: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/20.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 21: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/21.jpg)
Introduction Method Test Cases First Results Perspectives
Multi Crystal Data Collection
� Sample on mesh loop
� Mesh scan of sample
� Detection of protein diffraction
� Series of partial data collection
� Integration of partial sets
� Hierarchical cluster analysis
� Data merging
![Page 22: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/22.jpg)
Introduction Method Test Cases First Results Perspectives
Test Cases
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Introduction Method Test Cases First Results Perspectives
LCP Membrane Protein: Bacteriorhodopsin
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Introduction Method Test Cases First Results Perspectives
LCP Membrane Protein: Bacteriorhodopsin
LCP Crystallization Drop, crystalsize ca. 5 µm
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Introduction Method Test Cases First Results Perspectives
LCP Membrane Protein: Bacteriorhodopsin
LCP Crystallization Drop, crystalsize ca. 5 µm
Heat Map after Mesh Scan:59 spots picked for partial data
collection
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Introduction Method Test Cases First Results Perspectives
LCP Membrane Protein: Bacteriorhodopsin
hierarchical cluster analysis:10 out of 38 integrated datasets
selected for merging
Heat Map after Mesh Scan:59 spots picked for partial data
collection
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Introduction Method Test Cases First Results Perspectives
LCP Membrane Protein: Bacteriorhodopsin
Statistics:Space group: P 63Resolution: 19.73-2.57 (2.71-2.57)Rp.i.m.: 0.127 (0.546)Completeness: 97.7 (87.1)I/Sigma(I): 6.7 (1.8)Rwork: 0.18983Rfree: 0.20547
Bacteriorhodopsin:Biological Assembly
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Introduction Method Test Cases First Results Perspectives
LCP Membrane Protein: Bacteriorhodopsin
Statistics:Space group: P 63Resolution: 19.73-2.57 (2.71-2.57)Rp.i.m.: 0.127 (0.546)Completeness: 97.7 (87.1)I/Sigma(I): 6.7 (1.8)Rwork: 0.18983Rfree: 0.20547
Retinal Cofactor in Fo-Fc map(cutoff 2σ)
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Introduction Method Test Cases First Results Perspectives
Low Symmetry: Monoclinic Lysozyme
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Introduction Method Test Cases First Results Perspectives
Low Symmetry: Monoclinic Lysozyme
Crystallization Drop
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Introduction Method Test Cases First Results Perspectives
Low Symmetry: Monoclinic Lysozyme
Crystallization Drop Heat Map after Mesh Scan:54 spots picked for partial data
collection
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Introduction Method Test Cases First Results Perspectives
Low Symmetry: Monoclinic Lysozyme
hierarchical cluster analysis:20 out of 40 integrated datasets
selected for merging
Heat Map after Mesh Scan:54 spots picked for partial data
collection
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Introduction Method Test Cases First Results Perspectives
Low Symmetry: Monoclinic Lysozyme
Statistics:Space group: P21Resolution: 19.73-1.59 (1.68-1.59)Rp.i.m.: 0.080 (0.486)Completeness: 85.0 (82.1)I/Sigma(I): 8.0 (2.2)Rwork: 0.21291Rfree: 0.26489
mC Lysozyme: Secondary Structure
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Introduction Method Test Cases First Results Perspectives
Low Symmetry: Monoclinic Lysozyme
Statistics:Space group: P21Resolution: 19.73-1.59 (1.68-1.59)Rp.i.m.: 0.080 (0.486)Completeness: 85.0 (82.1)I/Sigma(I): 8.0 (2.2)Rwork: 0.21291Rfree: 0.26489
2Fo-Fc Map Example (cutoff 1 σ)
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Introduction Method Test Cases First Results Perspectives
Experimental Phasing: Thermolysin
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Introduction Method Test Cases First Results Perspectives
Experimental Phasing: Thermolysin
Crystals on Mesh Loop
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Introduction Method Test Cases First Results Perspectives
Experimental Phasing: Thermolysin
Crystals on Mesh Loop
Heat Map after Mesh Scan:96 spots picked for partial data
collection
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Introduction Method Test Cases First Results Perspectives
Experimental Phasing: Thermolysin
hierarchical cluster analysis:49 out of 77 integrated datasets
selected for merging
Heat Map after Mesh Scan:96 spots picked for partial data
collection
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Introduction Method Test Cases First Results Perspectives
Experimental Phasing: Thermolysin
ShelxD Results ShelxE Results
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Introduction Method Test Cases First Results Perspectives
Experimental Phasing: Thermolysin
Statistics:Space group: P 6122Resolution: 19.88-1.27 (1.33-1.27)Rp.i.m.: 0.017 (0.344)Completeness: 92.0 (53.4)I/Sigma(I): 25.4 (2.7)Rwork: 0.16989Rfree: 0.19703
Thermolysin Secondary Structure,Zn-Atom and Ca-Atoms in
anomalous map (4.5 σ)
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Introduction Method Test Cases First Results Perspectives
First Results
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Introduction Method Test Cases First Results Perspectives
Sanofi - AV
AV151
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Introduction Method Test Cases First Results Perspectives
Sanofi - AV
AV151
On-axis camera snapshot
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Introduction Method Test Cases First Results Perspectives
Sanofi - AV
AV151
On-axis camera snapshot
Mesh scan: 38 positions, 34 setsintegrated
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Introduction Method Test Cases First Results Perspectives
Sanofi - AV
AV56
On-axis camera snapshot
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Introduction Method Test Cases First Results Perspectives
Sanofi - AV
AV56
On-axis camera snapshot Mesh scan: 32 positions, 30 setsintegrated
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Introduction Method Test Cases First Results Perspectives
Sanofi - AV
Statistics
AV151 (19 sets):Resolution: 20.00-2.1 (2.16-2.10)Rp.i.m.: 0.064 (0.532)Completeness: 99.5(99.5)I/Sigma(I): 10.5 (2.4)
AV56 (19 sets):Resolution: 20.0-2.20 (2.27-2.00)Rp.i.m.: 0.086 (0.688)Completeness: 99.6 (99.9)I/Sigma(I): 9.5 (2.1)
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Introduction Method Test Cases First Results Perspectives
Perspectives
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Introduction Method Test Cases First Results Perspectives
Perspectives
� Characterization: Estimate Resolution
� Burn Strategy
� Room Temperature
� In situ
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Introduction Method Test Cases First Results Perspectives
Perspectives
� Characterization: Estimate Resolution X
� Burn Strategy
� Room Temperature
� In situ
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Introduction Method Test Cases First Results Perspectives
Perspectives
� Characterization: Estimate Resolution X
� Burn Strategy
� Room Temperature
� In situ
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Introduction Method Test Cases First Results Perspectives
Perspectives
� Characterization: Estimate Resolution X
� Burn Strategy
� Room Temperature
� In situ
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Introduction Method Test Cases First Results Perspectives
Perspectives
� Characterization: Estimate Resolution X
� Burn Strategy
� Room Temperature
� In situ
![Page 54: Multi Crystal Data Collection · 2015-03-13 · Introduction Method Test Cases First Results Perspectives Multi Crystal Data Collection A New Synchrotron Serial Crystallography Method](https://reader035.fdocuments.in/reader035/viewer/2022070810/5f095a097e708231d4266c3b/html5/thumbnails/54.jpg)
Introduction Method Test Cases First Results Perspectives
Fin.