[phenixbb] Autosol: MIRAS

Gabor Bunkoczi gb360 at cam.ac.uk
Thu Sep 6 02:52:00 PDT 2012


Dear AK,

if you just need the Se positions, you can run Phaser MR-SAD (available 
from the GUI) using your partial model as a starting point, together with 
the Se-Met data. Dependending on the model quality (and the model does not 
have to be very good), it may be able to locate the Se-sites.

BW, Gabor

On Sep 6 2012, ash.k at aol.com wrote:

> Satisfactory map means: I am expecting a coiled coil or helix bundle type 
> of assembly and I could see some densities appropriate for helices. There 
> are proper solvent channels and continuous stretches of densities.
>
> More to add about data on this: the data is anisotropic and the longer 
> 'c' axis and alignment of helical density along c axis support this. This 
> also makes me think that perhaps map is sensible.
>
> Few rounds of model building, refinement and DM has been successful to 
> assign around 10 polyala helices and their distribution looks sensible 
> from packing point of view. Now the problem is to assign the side chain. 
> I was hoping to make use of Se locations for this. R and Rfree are still 
> random, in the range of 50%. I am not sure, but it could be perhaps 
> because most of the scattering material is still unassigned.
> 
>
> 
>
> 
>
>-----Original Message-----
>From: Francis E Reyes <Francis.Reyes at colorado.edu>
>To: PHENIX user mailing list <phenixbb at phenix-online.org>
>Cc: phenixbb <phenixbb at phenix-online.org>
>Sent: Thu, Sep 6, 2012 8:07 am
>Subject: Re: [phenixbb] Autosol: MIRAS
>
>
> This one is solvable, but with extreme difficulty. I recently completed a 
> structure solution with experimental phases starting at 5.0 A using phase 
> information from multiple derivatives.
>
>
>How would you describe a somewhat satisfactory map?
>
>
>F
>
>On Sep 5, 2012, at 7:08 PM, ash.k at aol.com wrote:
>
>
>
>
>
> 
>
>Hi Shya,
>
> I did wavelength scan, got a good signal for Se and used appropriate 
> wavelengths for data collection and also used experimental f' and f'' 
> values for phasing. I think the reasons SAD or MAD for SeMet data is not 
> working are (i) low resolution: 3.7 for SeMet (Anomalous data is up to 
> 4.8A) (ii) I should have mentioned this earlier: 3 out of 6 Se are very 
> close to N-terminus, possible they are disordered. Unit cell is also some 
> what big..100, 120 and 320A; F222 space group.
>
>AK  
>
> 
>
>-----Original Message-----
>From: Shya Biswas <shyabiswas at gmail.com>
>To: PHENIX user mailing list <phenixbb at phenix-online.org>
>Sent: Thu, Sep 6, 2012 7:29 am
>Subject: Re: [phenixbb] Autosol: MIRAS
>
>
>Hi AK,
>Did you do a wavelength scan when you collected the SE dataset you
>need to put the values of f' and f'' from your wavelength scan in
>order to locate the heavy atom sites, 6 methionine should be enough to
>phase your molecule.
>Shya
>
>On Wed, Sep 5, 2012 at 9:25 PM,  <ash.k at aol.com> wrote:
>> Hi all,
>>
>> I am trying to solve a structure through experimental phasing using 
>> AUTOSOL. I have a couple of heavy atom derivative datasets (Hg, La, Eu, 
>> Cd) and also a SeMet data. Unfortunately all the datasets are of low 
>> resolution (3.7-4.2A) and there are possibly 4-8 molecules in the asu. 
>> MIR, SAD and MAD alone did not give any convincing solution.
>>
>> However, MIRAS, with a combination of few heavy atom datasets and the 
>> anomalous data from SeMet crystals, gave a somewhat satisfactory map. 
>> But the heavy atom site picked by AUTOSOL list only one of the heavy 
>> atoms i.e. Lanthanum. In another set of run, the solution of which was 
>> not convincing, the heavy atom substructure had only Hg. There are 6 Met 
>> out of 200 residues in one molecule and mass spec results show that Se 
>> incorporation is 100%.
>>
>> Now, my doubt is that why does the heavy atom substructure contain only 
>> La and how can I get the substructure involving Se from this solution 
>> (or the datasets used)? Se location is going to help me a lot for 
>> finding a starting point to assign side chains.
>>
>> Any suggestion would be greatly appreciated.
>>
>> Thanks
>> AK
>>
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     Dr Gabor Bunkoczi

     Cambridge Institute for Medical Research
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