I have used CNS before, but not for this sort of refinement. I see in
the bindividual.inp file that I can "select atoms to be included"--it
is defaulted at "known and not hydrogen". Do you know the proper
nomenclature for selecting a Zn ion in chains A and B?
Thanks,
Chris
On 5/6/14, Steven Herron <[log in to unmask]> wrote:
>
> Refining the occupancy will help your R-factor and flatten your density,
> but you need to be careful to also refine the B-factor of the metal
> ion. Don't refine both the occupancy and the B-factor during the same
> run (the two are correlated at this resolution), refine the occupancy of
> just the metal ion and then refine the B-factor of just the metal ion
> (repeat as needed). I used X-plor/CNS to do my refinements, so it was
> easy to refine the occupancy (or B-factor) of just the metal ion. After
> a few rounds of refinement both parameters will stop changing and you
> will have your answer. The final B-factor of the metal ion should be
> similar to the amino acid residues that are coordinated to it.
>
> Soaking in several different ion concentrations and collecting
> additional datasets is also a good idea (if you have the time). I did
> this type of experiment once before (see: JBC 278(14):12271-7. [ Apr 4,
> 2003]) (or: http://www.ncbi.nlm.nih.gov/pubmed/12540845). I soaked in
> several different Ca2+ ion concentrations and was able to determine the
> binding affinity for that calcium ion using crystallography.
>
> To make sure I was not stuck in a local minima, I would modify either
> the occupancy of the B-factor of the metal while keeping the other fixed
> and do a refinement. I even tried both large and small changes (both
> increases and decreases in value). It always came back to the earlier
> answer.
>
> Different Ca2+ ion concentrations can give some additional insight into
> the metal binding site. Between the no-Ca2+ structure and the high-Ca2+
> structure there was a conserved Asp-residue that changed conformation.
> So, I soaked in the appropriate amount of Ca2+ to see the residue in
> both positions. There was a high correlation between the asp residue
> orientation and the Ca2+ ion occupancy.
>
> Steven Herron
> [log in to unmask]
>
>
>
>
> On 5/6/2014 11:02 AM, Chris Fage wrote:
>> Hi Everyone,
>>
>> In my 2.5-angstrom structure, there is negative Fo-Fc density
>> surrounding a metal ion after refining in Phenix. From anomalous
>> diffraction I am certain of the metal's identity and position in each
>> monomer. Also, the ion is appropriately coordinated by nearby side
>> chains. Should I be refining the occupancy of the ion in attempt to
>> "flatten" the negative density? I am considering soaking the metal ion
>> into crystals or cocrystallizing and collecting additional datasets.
>>
>> Thanks for your help!
>>
>> Regards,
>> Chris
>
>
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