international conference on structural genomics 2006 hands-on workshop iii international conference...
TRANSCRIPT
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International Conference on Structural Genomics 2006
Hands-on Workshop III
International Conference on Structural Genomics 2006
Hands-on Workshop III
Automated Structural Determination by X-ray Crystallography
Automated Structural Determination by X-ray Crystallography
Yokohama, Japan October 19-20, 2006Yokohama, Japan
October 19-20, 2006
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OASIS-2006OASIS-2006Institute of Physics
Chinese Academy of SciencesBeijing 100080, P.R. China
Institute of PhysicsChinese Academy of Sciences
Beijing 100080, P.R. China
http://cryst.iphy.ac.cnhttp://cryst.iphy.ac.cn
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OASIS (2000)OASIS (2000) in CCP4in CCP4
OASIS-2004OASIS-2004 on the Webon the Web
OASIS-2006OASIS-2006 on the Web soonon the Web soon
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OASIS-2006 GUI for CCP4iOASIS-2006 GUI for CCP4i
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1. Direct-method
2. Reciprocal-space
1. Direct-method
2. Reciprocal-space
Functions of OASIS Functions of OASIS
SAD/SIR PhasingSAD/SIR Phasing
Fragment ExtensionFragment ExtensionFragment ExtensionFragment ExtensionDual-spaceDual-space
by combining with
DM, RESOLVE and ARP/wARPby combining with
DM, RESOLVE and ARP/wARP
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Why direct methods?Why direct methods?
For better initial SAD phases!
For better initial SAD phases!
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Bimodal distributionfrom SAD
" " "
The phase ofF”
P
Phase information available in SADPhase information available in SAD
Cochrandistribution
Peaked atany where
from 0 to 2
Peaked at
"2
Sim distribution
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Two different kinds of initial SAD phasesTwo different kinds of initial SAD phases
P+-modified phases
P++P
PSim PBimodal Sim-modified phases
P+
PSim PCochran
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" h h h1
" is the phase of " "exp 2N
j jj
i f i
h h h rF
1 1cos 2 " cos2 "
F h
h h h hh
F F FF
2 ' 'E E E h h h h 3 ' '" " " h h h h
1tan( ) 2 ( ) sin cos
2best P
h h h h
1
2 21 1exp 2 2 1 cos2 cos2
2 2m P
2h h h h h
,
2 2
2U T
pN N
j jj j
E E
Z Z
h h ' " h h h
, 3
1 1tanh sin
2 2
sin sin
c
best best
P
m m
h h
h' h h' h h' h' h h' hh'
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Comparison of 4 typical reflections from the protein histone methyltransferase SET 7/9
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Comparison of cumulative phase errors in descending order of Fobs
Comparison of cumulative phase errors in descending order of Fobs
60.263.415000
58.461.913500
56.960.812000
62.365.216352
55.659.410500
54.158.79000
52.957. 87500
51.257.06000
50.056.54500
49.157.13000
45.857.11500
Errors of PErrors of P++-modified -modified
phases phases (( oo ))Number of reflectionsNumber of reflections Errors of Sim-modified Errors of Sim-modified
phases phases (( oo ))
Histone methyltransferase SET 7/9
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Why dual-space?Why dual-space?
For less systematic errors!
For less systematic errors!
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Reciprocal-space fragment extension
OASIS + DM
Reciprocal-space fragment extension
OASIS + DM
Dual-space fragment extensionDual-space fragment extension
, 3
1 1tanh sin
2 2
s s inin
best best
P
m m
h h
h' h h' h h' h' hh h'h'
, 3
1 1tanh sin
2 2
s s inin
best best
P
m m
h h
h' h h' h h' h' hh h'h'
Real-spacefragment extension
RESOLVE BUILD and/or ARP/wARP
Real-spacefragment extension
RESOLVE BUILD and/or ARP/wARP
Partialstructure
Partialstructure
NoNo
YesYes
OK?OK?
EndEnd
PartialmodelPartialmodel
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OASIS ApplicationsOASIS Applications
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Ab initio SAD phasingAb initio SAD phasingwith
weak anomalous signalwith
weak anomalous signal
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SHARP-SOLOMON-ARP/wARPSHARP-SOLOMON-ARP/wARP
XylanaseSpace group: P21 Unit cell: a = 41.07, b = 67.14, c = 50.81Å = 113.5o Number of residues in the ASU: 303 Resolution limit: 1.75Å; Multiplicity: 15.9Anomalous scatterer: S (5 ) X-rays:synchrotron radiation= 1.488Å; f ” = 0.52Bijvoet ratio: <|F |>/<F > = 0.56% Data courtesy of Dr. Z. Dauter, National Cancer Institute, USA
OASIS-DM-RESOLVE BUILDcycle 025%
OASIS-DM-ARP/wARPcycle 699%SHARP-DM-RESOLVE BUILDBP3-DM-RESOLVE BUILD
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2.1Å 3.5Å 4.0Å
SAD phasing at different resolutionsTT0570 Cu-Kdata, <|F|>/<F> ~ 0.55%
SAD phasing at different resolutionsTT0570 Cu-Kdata, <|F|>/<F> ~ 0.55%
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OASIS + DM OASIS + DM
SOLVE/RESOLVE resultsSOLVE/RESOLVE results
improved byimproved by
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SOLVE/RESOLVE
SOLVE/RESOLVE +OASIS-DM-RESOLVE (BUILD)
Dual-space fragment extension based on SOLVE/RESOLVE resultsDual-space fragment extension based on SOLVE/RESOLVE results1LIA (d14) 2.8Å SIR data
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SOLVE/RESOLVE map
Sigma_A map based on a model manually builtfrom the SOLVE/RESOLVE map
OASIS-DM mapbased on
the same model
Dual-space fragment extension based on SOLVE/RESOLVE resultsDual-space fragment extension based on SOLVE/RESOLVE results2GW1 3.3Å SAD data
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Dual-space fragment extension based on SOLVE/RESOLVE resultsDual-space fragment extension based on SOLVE/RESOLVE results2GW1 3.3Å SAD data
Sigma_A map based on a model manually builtfrom the SOLVE/RESOLVE map
SOLVE/RESOLVE map OASIS-DM mapbased on
the same model
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Molecular Replacement Molecular Replacement
Fragment extensionFragment extension
based onbased on
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Fragment extension based on
molecular replacement
Finalmodel
acidic phospholipase A2124 residues
MRmodel
60 residues48 residues
ARP/wARP-OASIS-DM iteration
Cycle 1 Cycle 2 Cycle 3
ARP/wARP-DM iteration
Cycle 9 Cycle 11 Cycle 13
ARP/wARP-DM iteration
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A difficult case: 1UJZA difficult case: 1UJZSpace group: I 222Unit cell: a=62.88, b=74.55, c=120.44ÅNumber of residues in ASU: 215 molecule A: 87 residues molecule B: 128 residuesNumber of residues in the starting model: 46 (all in Mol. A)Resolution range: 37.57 – 2.10ÅNumber of reflections: 16460
Space group: I 222Unit cell: a=62.88, b=74.55, c=120.44ÅNumber of residues in ASU: 215 molecule A: 87 residues molecule B: 128 residuesNumber of residues in the starting model: 46 (all in Mol. A)Resolution range: 37.57 – 2.10ÅNumber of reflections: 16460
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Fragment ExtensionFragment ExtensionDual-spaceDual-space
without SAD/SIR informationwithout SAD/SIR information
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, 3
1 1tanh sin
2 2
sin si n
best best
P
m m
h h
h' h h' h h' h' hh h'h'
Partialstructure
Partialstructure
" h 5%
the phase of atoms
randomly selected from the current model
" h h h
" . . "model modeli e h h h h
Density modification
by DM
Density modification
by DM
NoNo
MRmodel
MRmodel
YesYes
EndEnd
Model rebuild by
RESOLVE BUILD or ARP/wARP
Model rebuild by
RESOLVE BUILD or ARP/wARP
OK?OK?
Phase improvement
by OASIS
Phase improvement
by OASIS
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P+ > 0.5
” model
P+ < 0.5
” model
<||> ~
<||> ~
”
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Range of phase error in degrees
Cycle 1 Cycle 3 Cycle 5 Cycle 7
Nr. of Reflns.
% of P+ > ½
Nr. of Reflns.
% of P+ > ½
Nr. of Reflns.
% of P+ > ½
Nr. of Reflns.
% of P+ > ½
0 - 30 3337 55 3761 60 4197 65 5733 76
30 - 60 2856 48 2874 49 3049 52 2469 43
60 - 90 2635 38 2359 38 2279 35 1852 18
90 - 120 2281 31 2189 27 1975 24 1702 7
120 -150 2175 24 2097 21 1858 16 1813 6
150 - 180 2064 22 2071 17 1991 13 1779 5
1UJZ Phase Statistics1UJZ Phase Statistics
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46 residues13 with side chains
MRmodelMRmodel
Cycle 2
ARP/wARP-DMiterationCycle 1
FinalmodelFinalmodel
215 residues
Cycle 1 Cycle 3
ARP/wARP-OASIS-DM iteration
Cycle 7Cycle 5201 residuesall with side chains
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Hai-fu Fan1, Yuan-xin Gu1, Tao Jiang2, Zheng-jiong Lin2 & Chao-de Zheng1
Hai-fu Fan1, Yuan-xin Gu1, Tao Jiang2, Zheng-jiong Lin2 & Chao-de Zheng1
Participants of this projectParticipants of this projectStaffsStaffs
Ph.D. studentsPh.D. studentsJian-rong Chen1, Qiang Chen3, Yao He1,
Sheng Huang1,4, He Li2, Jia-wei Wang1, Li-jie Wu1, De-qiang Yao1,5 & Tao Zhang1,6
Jian-rong Chen1, Qiang Chen3, Yao He1, Sheng Huang1,4, He Li2, Jia-wei Wang1, Li-jie Wu1,
De-qiang Yao1,5 & Tao Zhang1,6
1 Institute of Physics, CAS, Beijing, China2 Institute of Biophysics, CAS, Beijing, China3 Peking University, Beijing, China4 Institute of High Energy Physics, CAS, Beijing, China5 Univ. of Science & Technology of China, Hefei, China6 Lanzhou University, Lanzhou, China
1 Institute of Physics, CAS, Beijing, China2 Institute of Biophysics, CAS, Beijing, China3 Peking University, Beijing, China4 Institute of High Energy Physics, CAS, Beijing, China5 Univ. of Science & Technology of China, Hefei, China6 Lanzhou University, Lanzhou, China
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SAD data used in this presentationwere kindly provided by
SAD data used in this presentationwere kindly provided by
AcknowledgementsAcknowledgements
Dr. Z. Dauter1, Dr. S. J. Gamblin2, Dr. B. D. Sha3, Prof. I. Tanaka4, Dr. N. Watanabe4 & Dr. B. Xiao2
Dr. Z. Dauter1, Dr. S. J. Gamblin2, Dr. B. D. Sha3, Prof. I. Tanaka4, Dr. N. Watanabe4 & Dr. B. Xiao2
1 Argonne National Laboratory, USA2 The National Institute for Medical Research, UK3 Department of Cell Biology, University of Alabama at Birmingham, USA4 Graduate School of Science, Hokkaido University, Japan
1 Argonne National Laboratory, USA2 The National Institute for Medical Research, UK3 Department of Cell Biology, University of Alabama at Birmingham, USA4 Graduate School of Science, Hokkaido University, Japan
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Thank you!Thank you!