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Hi Wendy,
I had the same scenario a few years ago. It was zinc (from no obvious
source) and we identified/confirmed it by by X-ray fluorescence at a
synchrotron source
Cheers
J
Wendy Gordon wrote:
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Hello-
Thanks for entertaining my non-ccp4 question...
I am refining a crystal structure at ~2 Angstroms resolution in which I
find a large, unexpected electron density that is potentially a Zinc
ion because it is coordinated by 2 Histidines, 1 Glutamic acid, and
possibly a water molecule.
This site appears as a strong peak in anomalous difference Patterson
maps (the data was collected at SelMet wavelength 0.979 Angstroms and I
believe that Zn's absorption edge occurs around 1.2 Angstroms). The
problem is, I didn't add any zinc in any purification or
crystallization conditions. I DID affinity purify the protein with
nickel beads, so potentially it could be a Nickel ion. I should also
say that during refinement in refmac where my Zn occupancy is held at
1, that I obtain a negative peak in the 2Fo-Fc in this position, but if
I leave the site unoccupied- I get a huge positive peak- so I either
have the wrong species defined or my occupancy is not 100%- right?
Is there any way short of biochemical means (ITC, mutation, etc.) to
figure out what species is occupying this electron density? I have
thought of atomic absorption- has anyone tried it to determine the
metal species in a protein? Does it seem possible that I could have a
Zn ion in my protein crystal where the Zn could only come from our
standard DI water supply?
Thanks so much!
Wendy Ryan Gordon
--
Joel Tyndall, PhD
Lecturer
National School of Pharmacy
University of Otago
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