[3dem] Membrane protein structure: orientation issue

[Shashi Bhushan]

Dear All,

Does it mean that we can reconstruct a 3D map of a membrane protein having
C3 symmetry just from pure side views?

Best Wishes,

Shashi

On Thu, Mar 26, 2020 at 4:25 PM Christopher Aylett <chsaylett at gmail.com>
wrote:

> Hi Philip,
>
> This is why single particle analysis is a local minimisation dependent on
> the initial model used. The assigned orientations are based on this initial
> input, and providing Fourier space is well-sampled and the input is within
> the correct local minimum, the angular assignment and calculated 3D
> structure will converge.
>
> Best wishes,
>
> Chris
>
> ****
>
> > On 26 Mar 2020, at 07:44, Philip Köck <koeck at kth.se> wrote:
> >
> > Hi again.
> >
> > I believe you are right if you can be sure that you've found a single
> perfect side view and the symmetry is at least C3.
> > In that case I would say you can just symmetrize, as you describe, and
> get a rough 3D model without using either top or tilted views. Note: You
> are using a single side view!
> >
> > I don't see how a spread of different side views would help you though.
> > You run into the problem that Steve pointed out.
> > You don't know how the different side views are oriented with respect to
> each other unless you have at least one
> > top or tilted view to lock the relative orientations.
> >
> > All the best,
> >
> > Philip
> >
> >
> > Från: Dmitry Lyumkis <dlyumkis at salk.edu>
> > Skickat: den 25 mars 2020 17:41
> > Till: Philip Köck
> > Kopia: Jacopo Marino; 3dem at ncmir.ucsd.edu
> > Ämne: Re: [3dem] Membrane protein structure: orientation issue
> >
> > If it’s a twofold rotationally symmetric object, adhered to the
> air-water interface along a single “side" view, then a projection along,
> e.g., phi=0/theta=90 samples the same Fourier plane as the second
> projection along phi=180/theta=90. The reconstruction  will behave in a
> manner that is identical to one that is composed of exclusively “top” views
> of a rotationally symmetric object (or, more simply, an asymmetric particle
> with one preferential orientation, normalizing for the number of asymmetric
> units). Basically,  you’ll end up hyper-sampling around one plane in both
> cases, except that the planes will lie along distinct axes of the
> transform. Both are bad cases and, in the absence of other views, will lead
> to a bad reconstruction. To be clear: this is assuming that  there’s only
> one preferential orientation along the side view of a two-fold rotationally
> symmetric particle, and not more. If you have 3-fold rotational symmetry,
> and the sample is adhered to its “side” view, projections separated by
> phi=120° spacing will  lead to sampling of Fourier planes separated by
> phi=60° (due to symmetry). Effectively, you add two additional planes.
> Assuming you have enough particles, and there’s a bit of spread in the phi
> angle, in most practical cases, the reconstruction of a 3-fold
> rotationally symmetric object will be complete (or nearly so). The higher
> the rotational symmetry, obviously the better.
> >
> >
> > To answer the original question, adding top views for your membrane
> protein will be negligible in the reconstruction. Looks like you should
> already get a very nice map from the current data. Give it a try.
> >
> >
> > Best,
> >
> >
> > Dmitry
> >
> >
> >
> >
> >
> >
> >
> >
> > 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
> >
> >
> >
> >
> >
> >
> >
> >
> >
> > On Mar 25, 2020, at 12:06 AM, Philip Köck <koeck at kth.se> wrote:
> >
> >
> >
> >
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