Hi Eleanor,
I agree that experimental phases may be needed, although I may take the
lazy way out and just aggressively screen for a less problematic crystal
form.
Best regards,
Mark
Mark A. Wilson
Associate Professor
Department of Biochemistry/Redox Biology Center
University of Nebraska
N118 Beadle Center
1901 Vine Street
Lincoln, NE 68588
(402) 472-3626
[log in to unmask]
On 9/3/15 4:06 PM, "CCP4 bulletin board on behalf of Eleanor Dodson"
<[log in to unmask] on behalf of [log in to unmask]> wrote:
>How similar is this data set to the previously crystallised one?
>I think you said there is no translation vector in the previous one so
>they can't be very similar.
>Maybe it is time for experimental phases!
>Eleanor
>
>
>
>
>
>On 3 September 2015 at 21:47, Adrian Goldman
><[log in to unmask]> wrote:
>
>This would be my feeling too - one real 21, a twin axis and
>pseudosymmetry. The standard perfect storm.
>
>Sent from my iPhone
>
>> On 3 Sep 2015, at 21:07, Mark Wilson <[log in to unmask]> wrote:
>>
>> Hi Eleanor,
>> Yes, of course you are correct about the beta~90° requirement for
>>possible
>> twinning here-I was mistakenly thinking about pseudomerohedry in higher
>> symmetry space groups. The L plot looks like well-behaved data, with a
>> straight line that closely tracks the model untwinned one. Pointless,
>> provided with data integrated in P1, choses P212121 with high
>>confidence
>> (0.95-0-.99), which is similar to the results of analysis using xtriage
>>in
>> PHENIX. Unsymmetrized cell angles are within 0.02-0.05° of 90°.
>>Finally,
>> the protein we crystallized is (to-be-tested wrong protein scenario
>>aside)
>> identical to the previously crystallized one, and our unit cell axes are
>> the same within 5% or so.
>> Best regards,
>> Mark
>>
>> Mark A. Wilson
>> Associate Professor
>> Department of Biochemistry/Redox Biology Center
>> University of Nebraska
>> N118 Beadle Center
>> 1901 Vine Street
>> Lincoln, NE 68588
>> (402) 472-3626
>> [log in to unmask]
>>
>>
>>
>>
>>
>>
>>> On 9/3/15 2:45 PM, "Eleanor Dodson" <[log in to unmask]> wrote:
>>>
>>> Disorder almost always produces streaked spots, but I guess it isn't
>>> compulsory!
>>>
>>> By the way, you can have twinning in Monoclinic if B ~ 90. without
>>>having
>>> a = c.
>>>
>>>
>>> . What does the L plot look like? Have you used pointless which gives
>>>the
>>> CC for each symmetry operator separately - sometimes that shows say the
>>> 00 l axis is more 2-fold ish than the 0k0 axis..
>>>
>>>
>>>
>>> Eleanor
>>> PS - If the related protein fits into a similar cell with the same SG
>>>is
>>> yours bigger?
>>>
>>>
>>>
>>>
>>> On 3 September 2015 at 18:56, Mark Wilson
>>> <[log in to unmask]> wrote:
>>>
>>> Dear Remy,
>>> Indeed, I think you may be correct and we're pursuing this now. A
>>>perfect
>>> 0.5 lattice translocation along b in P212121 would (I think) result in
>>>the
>>> pathology we observe. We do not see zones of streaked reflections in
>>>the
>>> images, but my thinking is that if the lattice defect is coincident
>>>with a
>>> crystallographic translation operator, perhaps we wouldn't expect to.
>>> Best regards,
>>> Mark
>>>
>>> Mark A. Wilson
>>> Associate Professor
>>> Department of Biochemistry/Redox Biology Center
>>> University of Nebraska
>>> N118 Beadle Center
>>> 1901 Vine Street
>>> Lincoln, NE 68588
>>> (402) 472-3626 <tel:%28402%29%20472-3626> <tel:%28402%29%20472-3626>
>>> [log in to unmask]
>>>
>>>
>>>
>>>
>>>
>>>
>>>> On 9/3/15 12:49 PM, "Remy Loris" <[log in to unmask]> wrote:
>>>>
>>>> Dear Mark,
>>>>
>>>> My suspicion is that what you observe here is a lattice disorder,
>>>> possibly related to what is described in
>>>>
>>>> Jimin Wang, Satwik Kamtekar, Andrea J. Berman and Thomas A. Steitz
>>>> (2005) Correction of X-ray intensities from single crystals containing
>>>> lattice-translocation defects Acta Cryst D61, 67-74.
>>>>
>>>> If your unit cell is offset statistically by 0.5 in the b-direction,
>>>> this should provide such a strong non-origin peak as you observe. In
>>>>the
>>>> cases that have been described until now, this type of disorder also
>>>> involves zones of nice sharp reflections and other zones with more
>>>> streaky reflections. Do you see something similar?
>>>> In order to use such data, they have to be corrected as described in
>>>>the
>>>> paper above.
>>>>
>>>> Possibly, the crystals with the 10% non-origin peak also have the
>>>> disorder, but much less pronounced so that omitting the required
>>>> correction did not prevent structure determination and refinement
>>>> (similar to let say a small fraction of merohedral twinning that is
>>>> overlooked).
>>>>
>>>> Remy Loris
>>>> Vrije Universiteit Brussel and VIB
>>>>
>>>> DOI: 10.1107/S0907444904026721
>>>>
>>>>> On 03/09/15 18:13, George Sheldrick wrote:
>>>>> Dear Mark,
>>>>>
>>>>> Since your resolution is good enough, perhaps you should try to solve
>>>>> it ab initio with Arcimboldo Lite. This has already solved a number
>>>>>of
>>>>> structures that turned out to be unexpected.
>>>>>
>>>>> Best wishes, George
>>>>>
>>>>>
>>>>>> On 09/03/2015 05:44 PM, Mark Wilson wrote:
>>>>>> Hi Herman,
>>>>>> A fair point-the odds of "depressing coincidence" do seem to be
>>>>>> climbing!
>>>>>> We did inspect the deposited data for a similar peak and, while one
>>>>>>is
>>>>>> present, it is only ~10% of the origin and at a different location.
>>>>>> We'll
>>>>>> do some due diligence on our end by re-dissolving crystals and
>>>>>> performing
>>>>>> mass spec. As there seems to be some interest in this, I'll update
>>>>>> once
>>>>>> we've figured it out, even if it's an embarrassing case of wrong
>>>>>> protein,
>>>>>> same cell.
>>>>>> Best regards,
>>>>>> Mark
>>>>>>
>>>>>> Mark A. Wilson
>>>>>> Associate Professor
>>>>>> Department of Biochemistry/Redox Biology Center
>>>>>> University of Nebraska
>>>>>> N118 Beadle Center
>>>>>> 1901 Vine Street
>>>>>> Lincoln, NE 68588
>>>>>> (402) 472-3626
>>>>>> [log in to unmask]
>>>>>>
>>>>>>
>>>>>>
>>>>>>
>>>>>>
>>>>>>
>>>>>> On 9/3/15 10:25 AM, "CCP4 bulletin board on behalf of
>>>>>> [log in to unmask]"<[log in to unmask] on behalf of
>>>>>> [log in to unmask]> wrote:
>>>>>>
>>>>>>> Dear Mark,
>>>>>>>
>>>>>>> In this case you will have to apply Baysian statistics: given the
>>>>>>> prior:
>>>>>>> same protein, same space group same cell dimensions and molecular
>>>>>>> replacement fails completely, the likelihood of having some
>>>>>>> depressing
>>>>>>> coincidence somewhere is approaches 100%!
>>>>>>>
>>>>>>> What I would do in addition to excellent suggestions you already
>>>>>>> got, is
>>>>>>> to try to download the Fobs from the pdb for the structures with
>>>>>>>the
>>>>>>> same
>>>>>>> protein, space group and cell dimensions, and calculate pattersons
>>>>>>> with
>>>>>>> those. Sometimes strong peaks appear in pattersons for no obvious
>>>>>>> reasons.
>>>>>>> I would also consider statistical disorder, which will not show up
>>>>>>>in
>>>>>>> twinning statistics since in this case F's (including phases) are
>>>>>>> added
>>>>>>> instead of I's. Anyways, it will be an interesting puzzle to solve!
>>>>>>>
>>>>>>> Good luck,
>>>>>>> Herman
>>>>>>>
>>>>>>>
>>>>>>> -----Ursprüngliche Nachricht-----
>>>>>>> Von: CCP4 bulletin board [mailto:[log in to unmask]] Im Auftrag
>>>>>>> von
>>>>>>> Mark Wilson
>>>>>>> Gesendet: Donnerstag, 3. September 2015 02:06
>>>>>>> An: [log in to unmask]
>>>>>>> Betreff: Re: [ccp4bb] Translational NCS with one molecule in ASU
>>>>>>>
>>>>>>> Dear CCP4 Community,
>>>>>>> I've had a number of helpful responses (on- and off-list) that I
>>>>>>>will
>>>>>>> briefly summarize via response, including information that I
>>>>>>>probably
>>>>>>> should have included in the original post. Many have suggested a
>>>>>>> wrong
>>>>>>> space group, which I agree seems probable. MR was attempted in
>>>>>>> PHASER
>>>>>>> with all possible choices of space group for a primitive
>>>>>>>orthorhombic
>>>>>>> lattice, and in all cases failed with no rotation or translation
>>>>>>> peaks
>>>>>>> above a Z-score of 5.
>>>>>>> I've not yet tried monoclinic lattices and will, but this still
>>>>>>> wouldn't
>>>>>>> explain (to me anyway) an apparently impossible combination of
>>>>>>> translational NCS in P212121 with a cell that can't accommodate a
>>>>>>> second
>>>>>>> molecule unless twinning was also present, which may be the case
>>>>>>>(as
>>>>>>> Eleanor suggested). Others have asked about evidence of missed
>>>>>>>weak
>>>>>>> reflections indicating a larger true cell, which I looked for but
>>>>>>> didn't
>>>>>>> see in these images. The crystal that was used was mounted at room
>>>>>>> temperature, so there is no opportunity for cryo artifacts to have
>>>>>>> done
>>>>>>> something strange to the cell.
>>>>>>> Other suggestions included the presence of strong internal
>>>>>>> symmetry in
>>>>>>> the molecule, which is present, but as a pseudo-threefold, which
>>>>>>> seems
>>>>>>> incompatible with my NCS centering operation. One respondent
>>>>>>> suggested
>>>>>>> that we've crystallized the wrong molecule, which is something I
>>>>>>>also
>>>>>>> worried about a bit. Although possible, the space group and cell
>>>>>>> for our
>>>>>>> crystal are both as previously reported for this protein by another
>>>>>>> group, so it would be a depressing coincidence if we crystallized
>>>>>>>the
>>>>>>> wrong protein in the same cell. I'll be happy to update if/when we
>>>>>>> figure
>>>>>>> this out should it be of interest to the board. Thank you all for
>>>>>>> your
>>>>>>> thoughtful responses, which arrived in impressive number in the
>>>>>>>time
>>>>>>> it
>>>>>>> took me to drive home.
>>>>>>> Best regards,
>>>>>>> Mark
>>>>>>>
>>>>>>> Mark A. Wilson
>>>>>>> Associate Professor
>>>>>>> Department of Biochemistry/Redox Biology Center University of
>>>>>>> Nebraska
>>>>>>> N118 Beadle Center
>>>>>>> 1901 Vine Street
>>>>>>> Lincoln, NE 68588
>>>>>>> (402) 472-3626 <tel:%28402%29%20472-3626>
>>>>>>><tel:%28402%29%20472-3626>
>>>>>>> [log in to unmask]
>>>>>>>
>>>>>>>
>>>>>>>
>>>>>>>
>>>>>>>
>>>>>>> On 9/2/15 5:30 PM, "CCP4 bulletin board on behalf of Eleanor
>>>>>>>Dodson"
>>>>>>> <[log in to unmask] on behalf of
>>> [log in to unmask]> wrote:
>>>>>>>
>>>>>>>> Well - a translation of 0 0.5 0 would generate absences along b
>>>>>>>>so
>>>>>>>> that the SG could be P212121 or P 21 2 21Š
>>>>>>>>
>>>>>>>>
>>>>>>>> I would suspect twinning and a monoclinic SG .
>>>>>>>> Or as we found sadly - half the protein had disappeared in the
>>>>>>>> crystallisation trials..
>>>>>>>>
>>>>>>>>
>>>>>>>> But such a translation must mean you almost have a halved unit
>>>>>>>>cell?
>>>>>>>> Another way of saying there isn't enough room for your molecule..
>>>>>>>>
>>>>>>>>
>>>>>>>>
>>>>>>>> On 2 September 2015 at 22:38, Shane Caldwell
>>>>>>>> <[log in to unmask]> wrote:
>>>>>>>>
>>>>>>>> Are you certain it's actually P212121? One possibility is you're
>>>>>>>>at
>>>>>>>> lower symmetry and the Patterson peak corresponds to the NCS
>>>>>>>>between
>>>>>>>> particles that are almost-but-not-quite crystallographically
>>>>>>>> equivalent. In that case, MR probably wouldn't work. Does
>>>>>>>> searching in
>>>>>>>> P1 find anything?
>>>>>>>>
>>>>>>>> Shane Caldwell
>>>>>>>>
>>>>>>>> McGill University
>>>>>>>>
>>>>>>>>
>>>>>>>>
>>>>>>>>
>>>>>>>>
>>>>>>>> On Wed, Sep 2, 2015 at 5:19 PM, Mark Wilson<[log in to unmask]>
>>>>>>>> wrote:
>>>>>>>>
>>>>>>>> Dear CCP4 community,
>>>>>>>> I've encountered a curious problem with some recently collected
>>>>>>>> data.
>>>>>>>> I have a 1.8 Å resolution dataset that scales well in P212121 (log
>>>>>>>> file
>>>>>>>> available upon request). The unit cell parameters are similar to
>>>>>>>> those
>>>>>>>> reported for crystals of the same protein in a previous
>>>>>>>>publication,
>>>>>>>> although my crystallization condition is different. Nevertheless,
>>>>>>>> my
>>>>>>>> data produce a strong (47% of origin) peak in the Patterson map at
>>>>>>>> 0.0,
>>>>>>>> 0.5, 0.0, indicative of translational NCS. However, the unit cell
>>>>>>>> parameters can accommodate only one molecule in the ASU without
>>>>>>>> single-digit solvent content. Moreover, molecular replacement
>>>>>>>>with
>>>>>>>> a
>>>>>>>> model that should be nearly identical fails. Standard
>>>>>>>> intensity-based
>>>>>>>> tests show no evidence of twinning or other data pathology. Any
>>>>>>>> thoughts would be appreciated.
>>>>>>>> Best regards,
>>>>>>>> Mark
>>>>>>>>
>>>>>>>> Mark A. Wilson
>>>>>>>> Associate Professor
>>>>>>>> Department of Biochemistry/Redox Biology Center University of
>>>>>>>> Nebraska
>>>>>>>> N118 Beadle Center
>>>>>>>> 1901 Vine Street
>>>>>>>> Lincoln, NE 68588
>>>>>>>> (402) 472-3626<tel:%28402%29%20472-3626>
>>> [log in to unmask] <mailto:[log in to unmask]>
>>
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