[3dem] Abnormal FSC

[Wen Jiang]

We have encountered a few datasets with similar FSC oscillations and
we have identified beam tilt as the source of the problem after
refining/testing all particle parameters.

Wen Jiang, Ph.D.
Department of Biological Sciences
Department of Chemistry (Courtesy)
Purdue Cryo-EM Facility <http://cryoem.bio.purdue.edu>
Purdue University
Hockmeyer Hall of Structural Biology
240 S. Martin Jischke Drive
West Lafayette, IN 47907


On Tue, Aug 21, 2018 at 5:07 PM Andres Hernandez <aherna at scripps.edu> wrote:

> Hi 3DEM,
> We have been trying to diagnose the cause of this unusual FSC (sharp loss
> of correlation at ~4A) and have been unable to find the problem. This
> results in reported resolutions of ~4.3A when the map quality resembles
> that of similar ~3.5A reconstructions.
>
> We first thought that it could be due to a limited defocus range, but
> there is over 1.5 um of coverage in the particles for this reconstruction.
> I already tried local masks, spherical masks (different positions, radii),
> 3DFSCs, imposing symmetry, and many other things, but have been unable to
> identify the cause. The abnormal dip is present with both RELION and
> Cryosparc workflows.
>
> Any ideas what could be the cause? Has anyone seen an FSC like this
> before? We think it could be due to the molecule having multiple regions
> with pseudo-C7 symmetry or D7 symmetry. Imposing symmetry increases the
> correlation values beyond 4A (~2x) and reduces the dip slightly (~ 0.15
> FSC). However, we have gone through great lengths to minimize misalignment,
> yet localized reconstructions and localized FSCs still show this unusual
> feature.
>
>
> Thank you!
>
> Andres
> Lander Lab
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