*** For details on how to be removed from this list visit the *** *** CCP4 home page http://www.ccp4.ac.uk ***
It is entirely possible that you have zinc present. It is pretty ubiquitous and if bound strong enough can carry over from expression. Atomic absorption would tell you immediately which metal is present and is pretty straightforward to carry out. The coordination geometry will also give some clues to the potential identity of the metal ion. It is also possible that it is zinc but the site is not fully occupied. Mike > -----Original Message----- > From: [EMAIL PROTECTED] [mailto:[EMAIL PROTECTED] On > Behalf Of Wendy Gordon > Sent: Tuesday, May 09, 2006 4:30 PM > To: [email protected] > Subject: [ccp4bb]: Identity of unknown metal in protein crystal > > > *** 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 >
