[3dem] Membrane protein structure: orientation issue

[Dmitry Lyumkis]

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.

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<mailto:dlyumkis at salk.edu>
https://lyumkis.salk.edu






On Mar 24, 2020, at 10:53 PM, Jacopo Marino <jacopo.marino at psi.ch<mailto: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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--
Dr. Jacopo Marino
Laboratory of Biomolecular Research
Paul Scherrer Institute
OFLB/005
5232 Villigen PSI, Switzerland
tel: +41 56 310 5484 (or +41 56 310 5777)
e-mail: jacopo.marino at psi.ch<mailto:jacopo.marino at psi.ch>



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