From: ChenTiantian
Date: 14 October 2011 06:12
Hi there,
I am processing a dataset which has bad ice rings (as you can see in the attach png file).
I tried both XDS and imosflm, and got similar results, it seems that adding " EXCLUDE_RESOLUTION_RANGE" cannot get rid of the effects of the ice rings.
the following is part of the CORRECT.LP which is the second attached file, you can find more details there.
SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE >= -3.0 AS FUNCTION OF RESOLUTION
RESOLUTION NUMBER OF REFLECTIONS COMPLETENESS R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas Rmrgd-F Anomal SigAno Nano
LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr
4.24 37152 5537 5545 99.9% 46.9% 52.7% 37150 2.48 50.8% 19.4% -28% 0.513 5136
3.01 55344 9002 9840 91.5% 62.7% 65.1% 55116 1.76 68.3% 48.1% -28% 0.520 7760
2.46 84636 12699 12703 100.0% 67.4% 84.7% 84634 1.55 73.0% 54.2% -19% 0.513 12104
2.13 97910 14743 14987 98.4% 254.5% 199.3% 97908 0.16 276.2% 4899.9% -23% 0.473 14037
1.90 110260 16846 16940 99.4% 299.2% 303.3% 110245 0.06 325.0% -99.9% -17% 0.422 15995
1.74 118354 18629 18744 99.4% 1062.0% 1043.6% 118317 -0.20 1156.4% -99.9% -13% 0.380 17414
1.61 122958 20193 20331 99.3% 967.5% 1571.1% 122868 0.10 1059.7% 987.3% -2% 0.402 18348
1.51 125075 21554 21794 98.9% 838.9% 1355.1% 124933 0.08 922.6% 1116.9% -1% 0.402 18977
1.42 72057 17042 23233 73.4% 640.8% 775.3% 70391 0.08 732.5% 826.7% -8% 0.425 10003
total 823746 136245 144117 94.5% 166.4% 166.7% 821562 0.40 181.1% 296.7% -15% 0.435 119774
Note that I/SIGMA of each resolution shell is <2.5, so how should I do to process the dataset properly? Any suggestion about this super ice rings?
Thanks!
Tiantian
--
Shanghai Institute of Materia Medica, Chinese Academy of Sciences
Address: Room 101, 646 Songtao Road, Zhangjiang Hi-Tech Park,
Shanghai, 201203
----------
From: Stefan Gerhardt
try a frozen xtal ...
Albert-Ludwigs-Universität Freiburg
Inst.f.Org.Chem.u.Biochem
Albertstrasse 21
79104 Freiburg
Tel. +49 761 2035970
Fax. +49 761 2036161
----------
From: James Stroud
First of all, are you sure those are ice rings? They do not look typical. I think you might have salt crystals from dehydration *before* freezing. Otherwise, I think your freezing went well. Maybe try a humidity controlled environment when you freeze.
Second, I'm not so sure the bad stats come from the contaminating rings. The lattice seems to have some sort of problem, like a split lattice. You might be able to tackle this problem by increasing your spot size or skewing it's shape to compensate for the split. You need to investigate several images throughout the run to see whether and how to manipulate your spot size. Sometimes, the split lengthens the spots in the direction of the phi axis and you get lucky. But I think the phi axis might be horizontal in this picture, which makes things a little trickier. From one image, it is difficult to tell the pathology of this crystal.
In principle, if you can accurately measure the most high-resolution spots visible (which appear to be about 1.9 Å, guessing from your log file) then you will have a pretty good data set, even with the contaminating rings.
Personally, I'd use Denzo for this data, but I don't know what is vogue with the community right now. I still use O, so my tastes might be somewhat antiquated.
James
----------
From: Petri Kursula
----------
From: vandana kukshal
Hello ,
----------
From: Monolekha Bhattacharya
Hi Vandana,
The paper is attached herewith.
Bests,
monolekha
Monolekha Bhattacharya
Post Doctoral Fellow
Department of Chemistry and Biochemistry
University of Notre Dame
IN, USA
----------
From: James Holton
These rings are nanocrystalline cubic ice (ice Ic, as opposed to the "usual" ice Ih). It is an interesting substance in that noone has ever prepared a large single crystal of it. In fact, for very small crystals it can be hard to distinguish it from amorphous ice (or "glassy water"). The three main rings that you see from ice Ic coincide almost exactly with the centroids of the three main diffuse rings of glassy water, and as the ice Ic crystals get smaller, the rings get fatter (Scherrer broadening). You can even measure the size of the crystallites by measuring the width of the rings. At the limit of 1-2 unit cells wide, the diffraction pattern of ice Ic powder looks almost exactly like that of glassy water, so I suppose one could say that there is a continuum of phases between the two.
And yes, there are crystals that "like" a certain mixture of cubic ice and amorphous water in their solvent channels. Other's don't like it at all. But I agree with JS below that the problem here is not the ice rings. Probably overlaps? Best to look only at spots inside the 3.8A circle until you figure out what is going on.
-James Holton
MAD Scientist
----------
From: Kay Diederichs
Hi ChenTiantian,
the R-factors and I/sigma are bad even at low resolution where the first icering does not influence the results.
Thus, the problem with your data processing has little to do with the icerings. I guess that the indexing is not correct.
My suggestion:
1) using adxv or a similar display program, note what the inner and outer limits of the ice rings are. These values should be used as parameters for the EXCLUDE_RESOLUTION_RANGE= keywords in XDS.INP, not the provided ones (which are meant for hexagonal ice).
2) start XDS from the INIT step
3) use at least half of your DATA_RANGE as SPOT_RANGE
4) make sure that ORGX and ORGY are correct - mis-indexing is in 90% of the cases due to a wrong origin. In fact, just estimating ORGX and ORGY from the first frame, using adxv or XDS-viewer, seems to do a good job.
HTH,
Kay
Am 20:59, schrieb ChenTiantian:
----------
From: Harry
Hi
I'd agree with Kay here - I would think that the original indexing is incorrect.
One thing I notice on the original image as posted - there's a red cross on it - if that's supposed to mark the beam position, I think it's about 4mm or so away from the true position.
So -
(1) check the beam position carefully (it may be wrong in the image header)
(2) after indexing, make sure that the predictions match the spot positions
(3) if the predictions don't match the spot positions, don't try to integrate - find out what's wrong (wrong wavelength, beam position, distance???). If you can't work it out, ask one of the experts to look at a sample of your original images (iMosflm ask Andrew or me, XDS ask Kay, HKL Wladek or ZO...).
(4) If the predictions do match the spot positions, integrate the dataset in P1 (i.e. triclinic) and see what Pointless suggests as the symmetry. You may just be trying to impose too much symmetry. If you can't work out what the issue is, ask an expert to help directly - we're all happy to help out!
(5) Worry about the ice rings after you've sorted out the above problems, not before.
HTH
--
Dr Harry Powell, MRC Laboratory of Molecular Biology, MRC Centre, Hills Road, Cambridge, CB2 0QH
----------
From: Nian Huang
Hi,
I agree with other people. You must have a wrong index here. Can you tell us what is the unit cell for this crystal from your determination? I can see very close spots in the high resolution shell from your image, which are overlapped into one spot in the low resolution shell. Try to use other frames to do the indexing. If it is still not working, it might be easier to collect another dataset with better crystal alignment.
Best,
Nian
----------
From: Sanishvili, Ruslan
The silver lining of having ice (or any other kind) rings is that one can determine the origin rather accurately by just finding the center of these rings.
Cheers,
N.
Ruslan Sanishvili (Nukri), Ph.D.
GM/CA-CAT
Biosciences Division, ANL
9700 S. Cass Ave.
Argonne, IL 60439
----------
From: ChenTiantian
Hi there,
Thank you for all your suggestions and generous help, I tried some methods you guys mentioned and learned something new . I really appreciate it.
With Kay's help,(after exclusion of the ice rings he found that the data are P1, not P2(1). ) I got my structure solved, there are four copies in the AU,
I cut the high resolution to 2.4, and now the R/Rfree is 0.2254/0.2648, this is not the final result. I'm still working on it.
Thank you so much.
Best Regards,
Tiantian
Date: 14 October 2011 06:12
Hi there,
I am processing a dataset which has bad ice rings (as you can see in the attach png file).
I tried both XDS and imosflm, and got similar results, it seems that adding " EXCLUDE_RESOLUTION_RANGE" cannot get rid of the effects of the ice rings.
the following is part of the CORRECT.LP which is the second attached file, you can find more details there.
SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE >= -3.0 AS FUNCTION OF RESOLUTION
RESOLUTION NUMBER OF REFLECTIONS COMPLETENESS R-FACTOR R-FACTOR COMPARED I/SIGMA R-meas Rmrgd-F Anomal SigAno Nano
LIMIT OBSERVED UNIQUE POSSIBLE OF DATA observed expected Corr
4.24 37152 5537 5545 99.9% 46.9% 52.7% 37150 2.48 50.8% 19.4% -28% 0.513 5136
3.01 55344 9002 9840 91.5% 62.7% 65.1% 55116 1.76 68.3% 48.1% -28% 0.520 7760
2.46 84636 12699 12703 100.0% 67.4% 84.7% 84634 1.55 73.0% 54.2% -19% 0.513 12104
2.13 97910 14743 14987 98.4% 254.5% 199.3% 97908 0.16 276.2% 4899.9% -23% 0.473 14037
1.90 110260 16846 16940 99.4% 299.2% 303.3% 110245 0.06 325.0% -99.9% -17% 0.422 15995
1.74 118354 18629 18744 99.4% 1062.0% 1043.6% 118317 -0.20 1156.4% -99.9% -13% 0.380 17414
1.61 122958 20193 20331 99.3% 967.5% 1571.1% 122868 0.10 1059.7% 987.3% -2% 0.402 18348
1.51 125075 21554 21794 98.9% 838.9% 1355.1% 124933 0.08 922.6% 1116.9% -1% 0.402 18977
1.42 72057 17042 23233 73.4% 640.8% 775.3% 70391 0.08 732.5% 826.7% -8% 0.425 10003
total 823746 136245 144117 94.5% 166.4% 166.7% 821562 0.40 181.1% 296.7% -15% 0.435 119774
Note that I/SIGMA of each resolution shell is <2.5, so how should I do to process the dataset properly? Any suggestion about this super ice rings?
Thanks!
Tiantian
--
Shanghai Institute of Materia Medica, Chinese Academy of Sciences
Address: Room 101, 646 Songtao Road, Zhangjiang Hi-Tech Park,
Shanghai, 201203
----------
From: Stefan Gerhardt
try a frozen xtal ...
Dr Stefan Gerhardt
Albert-Ludwigs-Universität Freiburg
Inst.f.Org.Chem.u.Biochem
Albertstrasse 21
79104 Freiburg
Tel. +49 761 2035970
Fax. +49 761 2036161
----------
From: James Stroud
First of all, are you sure those are ice rings? They do not look typical. I think you might have salt crystals from dehydration *before* freezing. Otherwise, I think your freezing went well. Maybe try a humidity controlled environment when you freeze.
Second, I'm not so sure the bad stats come from the contaminating rings. The lattice seems to have some sort of problem, like a split lattice. You might be able to tackle this problem by increasing your spot size or skewing it's shape to compensate for the split. You need to investigate several images throughout the run to see whether and how to manipulate your spot size. Sometimes, the split lengthens the spots in the direction of the phi axis and you get lucky. But I think the phi axis might be horizontal in this picture, which makes things a little trickier. From one image, it is difficult to tell the pathology of this crystal.
In principle, if you can accurately measure the most high-resolution spots visible (which appear to be about 1.9 Å, guessing from your log file) then you will have a pretty good data set, even with the contaminating rings.
Personally, I'd use Denzo for this data, but I don't know what is vogue with the community right now. I still use O, so my tastes might be somewhat antiquated.
James
----------
From: Petri Kursula
Your main problem is not the ice rings but a wrong lattice/indexing solution. R factors are very high for even low res shells and I/sigma very low. To me this tells you are not finding your diffraction spots at all.
---
First thing to try: Take more images for the indexing step and use only the strongest spots. And do not refine distance during indexing, as you probably have a pretty high mosaicity.
Petri
---
Petri Kursula, PhD
Group Leader, Docent of Neurobiochemistry
Department of Biochemistry, University of Oulu, Finland
Department of Chemistry, University of Hamburg, Germany
Visiting Scientist (CSSB-HZI, DESY, Hamburg, Germany)
---
----------
From: vandana kukshal
Hello ,
Can any one send me pdf of this paper.... as its a old paper and not accessible here .
Vandana kukshal
M.F. Perutz, Preparation of haemoglobin crystals. J. Cryst. Growth, 2 (1968), pp. 54–56.
--
Vandana kukshal
----------
From: Monolekha Bhattacharya
Hi Vandana,
The paper is attached herewith.
Bests,
monolekha
Monolekha Bhattacharya
Post Doctoral Fellow
Department of Chemistry and Biochemistry
University of Notre Dame
IN, USA
----------
From: James Holton
These rings are nanocrystalline cubic ice (ice Ic, as opposed to the "usual" ice Ih). It is an interesting substance in that noone has ever prepared a large single crystal of it. In fact, for very small crystals it can be hard to distinguish it from amorphous ice (or "glassy water"). The three main rings that you see from ice Ic coincide almost exactly with the centroids of the three main diffuse rings of glassy water, and as the ice Ic crystals get smaller, the rings get fatter (Scherrer broadening). You can even measure the size of the crystallites by measuring the width of the rings. At the limit of 1-2 unit cells wide, the diffraction pattern of ice Ic powder looks almost exactly like that of glassy water, so I suppose one could say that there is a continuum of phases between the two.
And yes, there are crystals that "like" a certain mixture of cubic ice and amorphous water in their solvent channels. Other's don't like it at all. But I agree with JS below that the problem here is not the ice rings. Probably overlaps? Best to look only at spots inside the 3.8A circle until you figure out what is going on.
-James Holton
MAD Scientist
----------
From: Kay Diederichs
Hi ChenTiantian,
the R-factors and I/sigma are bad even at low resolution where the first icering does not influence the results.
Thus, the problem with your data processing has little to do with the icerings. I guess that the indexing is not correct.
My suggestion:
1) using adxv or a similar display program, note what the inner and outer limits of the ice rings are. These values should be used as parameters for the EXCLUDE_RESOLUTION_RANGE= keywords in XDS.INP, not the provided ones (which are meant for hexagonal ice).
2) start XDS from the INIT step
3) use at least half of your DATA_RANGE as SPOT_RANGE
4) make sure that ORGX and ORGY are correct - mis-indexing is in 90% of the cases due to a wrong origin. In fact, just estimating ORGX and ORGY from the first frame, using adxv or XDS-viewer, seems to do a good job.
HTH,
Kay
Am 20:59, schrieb ChenTiantian:
--
----------
From: Harry
Hi
I'd agree with Kay here - I would think that the original indexing is incorrect.
One thing I notice on the original image as posted - there's a red cross on it - if that's supposed to mark the beam position, I think it's about 4mm or so away from the true position.
So -
(1) check the beam position carefully (it may be wrong in the image header)
(2) after indexing, make sure that the predictions match the spot positions
(3) if the predictions don't match the spot positions, don't try to integrate - find out what's wrong (wrong wavelength, beam position, distance???). If you can't work it out, ask one of the experts to look at a sample of your original images (iMosflm ask Andrew or me, XDS ask Kay, HKL Wladek or ZO...).
(4) If the predictions do match the spot positions, integrate the dataset in P1 (i.e. triclinic) and see what Pointless suggests as the symmetry. You may just be trying to impose too much symmetry. If you can't work out what the issue is, ask an expert to help directly - we're all happy to help out!
(5) Worry about the ice rings after you've sorted out the above problems, not before.
HTH
Harry
--
Dr Harry Powell, MRC Laboratory of Molecular Biology, MRC Centre, Hills Road, Cambridge, CB2 0QH
----------
From: Nian Huang
Hi,
I agree with other people. You must have a wrong index here. Can you tell us what is the unit cell for this crystal from your determination? I can see very close spots in the high resolution shell from your image, which are overlapped into one spot in the low resolution shell. Try to use other frames to do the indexing. If it is still not working, it might be easier to collect another dataset with better crystal alignment.
Best,
Nian
----------
From: Sanishvili, Ruslan
The silver lining of having ice (or any other kind) rings is that one can determine the origin rather accurately by just finding the center of these rings.
Cheers,
N.
Ruslan Sanishvili (Nukri), Ph.D.
GM/CA-CAT
Biosciences Division, ANL
9700 S. Cass Ave.
Argonne, IL 60439
----------
From: ChenTiantian
Hi there,
Thank you for all your suggestions and generous help, I tried some methods you guys mentioned and learned something new . I really appreciate it.
With Kay's help,(after exclusion of the ice rings he found that the data are P1, not P2(1). ) I got my structure solved, there are four copies in the AU,
I cut the high resolution to 2.4, and now the R/Rfree is 0.2254/0.2648, this is not the final result. I'm still working on it.
Thank you so much.
Best Regards,
Tiantian
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