On Tuesday, October 30, 2012 01:44:43 pm Adrian Goldman wrote:
> The coordination is indicative but not conclusive but, as I responded to the original poster, I think the best approach is to use anomalous scattering. You can measure just below and above the Ca edge,
Actually, you can't. The Ca K-edge is at 3.07Å, which is not a wavelength
amenable to macromolecular data collection.
cheers,
Ethan
> and similarly with the Zn, and those maps will be _highly_ indicative of the relative amounts of metal ion present. In fact, you can deconvolute so that you know the occupancy of the metals at the various sites.
>
> Adrian
>
>
> On 30 Oct 2012, at 22:37, Chittaranjan Das wrote:
>
> > Veerendra,
> >
> > You can rule out if zinc has replaced calcium ions (although I agree with Nat and others that looking at the coordination sphere should give a big clue) by taking a few crystals, washing them a couple of times and subjecting them to ICP-MS analysis, if you have access to this technique. You can learn how many zinc, if any, have bound per one protein molecule in the dissolved crystal.
> >
> > Best
> > Chitta
> >
> >
> >
> > ----- Original Message -----
> > From: "Veerendra Kumar" <[log in to unmask]>
> > To: [log in to unmask]
> > Sent: Tuesday, October 30, 2012 2:55:33 PM
> > Subject: [ccp4bb] Ca or Zn
> >
> > Dear CCP4bb users,
> >
> > I am working on a Ca2+ binding protein. it has 4-5 ca2+ binding sites. I purified the protein in presence of Ca2+ and crystallized the Ca2+ bound protein. I got crystal and solved the structure by SAD phasing at 2.1A resolution. I can see the clear density in the difference map for metals at the expected binding sites. However I had ZnCl2 in the crystallization conditions. Now i am not sure whether the observed density is for Ca or Zn or how many of them are ca or zn? Since Ca (20 elctron) and Zn (30 electron), is this value difference can be used to make a guess about different ions?
> > is there any way we can find the electron density value at different peaks?
> >
> > Thank you
> >
> > Veerendra
>
--
Ethan A Merritt
Biomolecular Structure Center, K-428 Health Sciences Bldg
University of Washington, Seattle 98195-7742
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