[3dem] Membrane protein structure: orientation issue

[Basil Greber]

I am also confused by this. Shouldn’t a tomographic series (180 degrees worth of side views) do it for a C1 particle?

Basil



> Am 25.03.2020 um 00:06 schrieb Philip Köck <koeck at kth.se>:
> 
> ​Why do you say that the symmetry has to be at least C3?
> 
> All the best,
> 
> Philip
> 
> 
> 
> Från: 3dem <3dem-bounces at ncmir.ucsd.edu> för Dmitry Lyumkis <dlyumkis at salk.edu>
> Skickat: den 25 mars 2020 07:07
> Till: Jacopo Marino
> Kopia: 3dem at ncmir.ucsd.edu
> Ämne: Re: [3dem] Membrane protein structure: orientation issue
>   
> Assuming this is a symmetric membrane protein with at least 3-fold rotational symmetry, you don’t need the top and bottom views to fully sample Fourier space and arrive at a high-quality reconstruction. Side-views of a rotationally symmetric particle are  sufficient, and the reconstruction will be complete. 
> 
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> 
> Dmitry Lyumkis
> Assistant Professor
> The Salk Institute for Biological Studies
> 10010 North Torrey Pines Road, La Jolla, CA 92037
> T: 858-453-4100 x1155
> E: dlyumkis at salk.edu
> https://lyumkis.salk.edu
> 
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> On Mar 24, 2020, at 10:53 PM, Jacopo Marino <jacopo.marino at psi.ch> wrote:
> 
> 
> How much do you really need top and bottom views for 3D reconstruction ? Have you tried ?
> 
> 
> On 25.03.2020 06:48, Saumya Verma Bajaj wrote:
> 
> 
> Dear all, 
> 
> I am trying to solve the structure of a tetrameric membrane protein complex, with the protein embedded in detergent micelle (0.05% GDN) and a soluble accessory protein attached to it.  
> 
> Following 2D classification, while the side views and oblique views are easily visible, and the top/bottom are very few (~1-2%) (please see attached image) and get further diminished in subsequent rounds of 2D classification.     
> 
> While we are trying different grid types to overcome the orientation problem at the sample level, I was wondering if there are certain tweaks we can make to the analysis parameters (particle picking, box size, mask, ctf  etc.) to enhance the signal of protein embedded within a micelle in the current data set. We are using Relion3.1 for SPA. 
> 
> Any suggestions to salvage this set of data will be very helpful. 
> 
> Thank you.
> Best regards,
> Saumya 
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