To further clarify things, the data was collected at a synchrotron beamline
with collimator size ~130*40(um*square), beam divergence ~0.3*0.1mRad. The
detector type was MarCCD.
The crystal was multiple-faced trigonal (space group P3121) the size was
about 0.1*0.1*0.15mm. The exposure time was 2s for each image.
I am currently refining the structure, however the Rfree stays above 30%. A
close inspection shows at high resolution shell the spots become rod shaped.
As I said we are preparing new constructs with N-terminal his-tag cleaved.
But any other good suggestions out there might be helpful to avoid future
frustration.
Thanks,
Hubing
On Wed, Nov 24, 2010 at 10:26 PM, <[log in to unmask]> wrote:
> I think there may be two effects going on here:
>
>
>
> I think the “ears” on the round spots which also feature on the more rod
> shaped spots if you look closely could be related to a misalignment of the
> beamline optics.
>
>
>
> I think the change in spot shape from round to rod shaped is due to the
> crystal quality.
>
>
>
> Do the “ears” only feature on this image of this crystal or do they appear
> on other images? If the ear effect is a one off then that would tend to
> suggest it isn’t a beamline optic effect.
>
>
>
> Liz
>
>
>
> Dr. Liz Duke
>
> Principal Beamline Scientist
>
> Diamond Light Source
>
> Harwell Science and Innovation Campus
>
> Chilton
>
> OX11 0DE
>
> UK
>
>
>
> Tel. 01235 778057
>
> Mob. 07920 138148
>
>
>
> *From:* CCP4 bulletin board [mailto:[log in to unmask]] *On Behalf Of *Hubing
> Lou
> *Sent:* 24 November 2010 14:09
> *To:* [log in to unmask]
> *Subject:* [ccp4bb] unusual diffraction spots
>
>
>
> Dear CCP4BBer,
>
> I recently collected a dataset at synchrotron. The diffraction was quite
> anisotropic with one direction to 2.1Angstrom while the other is 3.0Ang.
> What unusual is in the diffraction image (see the attached file), clearly at
> low resolution there were some spots with tails ("two ears") and at the high
> resolution shell the spots turned to be rod-shaped. Please, can anyone
> explain how this could be? Is this related to the anisotropy? The protein
> was N-terminal his6-tagged, we are currently preparing new samples with
> the His-tag removed. But any other suggestions are also very welcomed.
>
> Regards,
>
> Hubing
>
>
>
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