*** For details on how to be removed from this list visit the *** *** CCP4 home page http://www.ccp4.ac.uk ***
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
