[phenixbb] Autosol: MIRAS

[ash.k at aol.com]

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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