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This question is very easily answered by doing a anomalous fourier with your calculated phases. CA has a significant anomalous signal at usual - Xray wavelengths and Mg does not. Eleanor On 7 March 2018 at 02:43, Rajesh Kumar <[log in to unmask]> wrote: > Hi Chris, > > I tried to refine with Ca++ but density became highly negative. I could > only refine it with Mg++ and Na+ without any green or red density. Na+ is > less likely because Na+-water distance normally 2.7 - 2.8 Å. The mg++ does > not have any interaction with the amino acid residues directly but waters > coordinated with Mg++ have hydrogen bonding interaction with protein > residues. > I have to analyze structure more carefully and come up with an explanation. > > -Rajesh > > > > > On Tue, Mar 6, 2018 at 8:37 PM, Christopher Squire < > [log in to unmask]> wrote: > >> At 200 mM CaCl2 (I think you said) I'd be surprised if this was not >> calcium. The distances compared to a dictionary are not always perfect. Put >> both a Mg(H2O)6 and a Ca(H2O)6 in place and refine. The temperature factors >> and/or residual electron density in maps will likely tell you which is the >> case. We have similar issues in some of our structures where the same site >> can bind both Ca and Mg but the crystallisation is in high concentrations >> of Ca so most likely calcium not magnesium. Otherwise, if the site is a >> very specific and strong magnesium binding site then it is possible that Mg >> is retained from earlier in your purification or even carried over from the >> bacterial expression medium. >> >> >> Chris. >> >> >> *Dr. Christopher Squire* >> Associate Professor, School of Biological Sciences >> University of Auckland >> Ph: +64 9 923 3086 <+64%209-923%203086> >> Ph: internal ext 83086 >> >> ------------------------------ >> *From:* CCP4 bulletin board <[log in to unmask]> on behalf of Rajesh >> Kumar <[log in to unmask]> >> *Sent:* 07 March 2018 1:07 PM >> *To:* [log in to unmask] >> *Subject:* Re: [ccp4bb] Unknown density >> >> Dear All, >> >> It is mg++ with 6 waters. As mg-water distances are 2 - 2.1 Å. Now from >> where I got the mg++ in the structure? We have not used Mg++ in >> purification and no Mg++ in the crystallization condition. >> >> But the inhibitor, which I used for the co-crystallization might have >> mg++. Because during drug synthesis, very often we use MgSO4 in drying >> steps. It might be coming from there. >> >> Thank you very much all, >> Rajesh >> >> >> >> >> >> >> On Tue, Mar 6, 2018 at 6:57 PM, Prem Prakash <[log in to unmask]> wrote: >> >>> Hi Rajesh, >>> >>> It might be water molecules present at the symmetry axis, one more water >>> molecule at the centre (exactly at the axis, with occupancy of 0.5). If you >>> put three water molecule you probably may find the other three >>> symmetry-mates water molecules in opposite sides. OR One water molecule at >>> the symmetry axis.surrounded by 5 different water molecules. Or You can try >>> As Garib mentioned it may be a six anions (or water) coordinated with some >>> metals possibly in crystallization or protein buffer. Please try refining >>> both cases parallely. I once encountered such case in my structure. >>> Probably it may help. >>> >>> Good luck >>> >>> Prem >>> >>> On Wed, Mar 7, 2018 at 4:37 AM, Abhik Mukhopadhyay < >>> [log in to unmask]> wrote: >>> >>>> Hi Rajesh, >>>> >>>> You may find this database is also useful while refining your model >>>> >>>> http://mespeus.bch.ed.ac.uk/MESPEUS_10/ >>>> >>>> and for calcium >>>> >>>> http://mespeus.bch.ed.ac.uk/MESPEUS_10/_5.jsp >>>> >>>> Abhik >>>> >>>> On Tue, Mar 6, 2018 at 10:59 PM, Patrick Loll <[log in to unmask]> wrote: >>>> >>>>> Calcium likes to form octahedral complexes with water (or other >>>>> oxygen-containing) ligands. This looks like a classic example. >>>>> >>>>> After you model and refine this, you’ll want to check water-metal >>>>> distances, to make sure they are appropriate for calcium. There is a nice >>>>> literature on such things, which I of course don’t have at my fingertips; >>>>> but I think Wladek Minor has done some data-mining in metal-containing >>>>> protein structures, and Amy Katz and Jenny Glusker have a number of papers >>>>> that are relevant. There are more, of course—a little time in the “library” >>>>> is warranted. >>>>> >>>>> Cheers, >>>>> >>>>> Pat Loll >>>>> >>>>> > On 6 Mar 2018, at 5:19 PM, Rajesh Kumar <[log in to unmask]> >>>>> wrote: >>>>> > >>>>> > Dear All, >>>>> > >>>>> > Have you had experience with this kind of density? I am wandering >>>>> what this could be? >>>>> > >>>>> > Thank you very much for the help. >>>>> > >>>>> > -Rajesh >>>>> > >>>>> > >>>>> > <Screen Shot 2018-03-06 at 5.15.20 PM.png> >>>>> > >>>>> >>>>> ------------------------------------------------------------ >>>>> --------------------------- >>>>> Patrick J. Loll, Ph. D. >>>>> Professor of Biochemistry & Molecular Biology >>>>> Drexel University College of Medicine >>>>> Room 10-102 New College Building >>>>> 245 N. 15th St >>>>> <https://maps.google.com/?q=245+N.+15th+St&entry=gmail&source=g>., >>>>> Mailstop 497 >>>>> Philadelphia, PA 19102-1192 USA >>>>> >>>>> (215) 762-7706 >>>>> [log in to unmask] >>>>> [log in to unmask] >>>>> >>>> >>>> >>> >>> >>> -- >>> With kind regards, >>> >>> Prem Prakash >>> PhD Research Scholar >>> Protein Crystallography Lab >>> Biosciences and Bioengineering >>> IIT Bombay >>> >> >> >