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, <elizabeth.d...@diamond.ac.uk> 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:ccp...@jiscmail.ac.uk] *On Behalf Of > *Hubing > Lou > *Sent:* 24 November 2010 14:09 > *To:* CCP4BB@JISCMAIL.AC.UK > *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 > > > > -- > > This e-mail and any attachments may contain confidential, copyright and or > privileged material, and are for the use of the intended addressee only. If > you are not the intended addressee or an authorised recipient of the > addressee please notify us of receipt by returning the e-mail and do not > use, copy, retain, distribute or disclose the information in or attached to > the e-mail. > Any opinions expressed within this e-mail are those of the individual and > not necessarily of Diamond Light Source Ltd. > Diamond Light Source Ltd. cannot guarantee that this e-mail or any > attachments are free from viruses and we cannot accept liability for any > damage which you may sustain as a result of software viruses which may be > transmitted in or with the message. > Diamond Light Source Limited (company no. 4375679). Registered in England > and Wales with its registered office at Diamond House, Harwell Science and > Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom > > >
