[phenixbb] Refine pixel size of map (for EM data)?

[Oliver Clarke]

Hi Reza,

Second question first - not much (I can't detect any), assuming same scope
and mag (which is relieving for multiple reasons). However, I don't know
how close the pixel size I obtain by real space correlation with a crystal
structure is to the "true" pixel size, and whether it is wrong in any
systematic way.

Re the other questions, I have not compared rigorously (I only refined
against the map with the correct pixel size), but I undertook steps to
correct it when I noticed during initial building in Coot that known
(solved) domains were not quite fitting the density as well as expected. I
would expect some consequences from this, particularly in terms of clash
analysis and the use of reference restraints (not sure about electrostatics
etc).

Where it does make a big difference is in comparing structures of the same
or related proteins solved by different groups - if the pixel size varies
by even 1-2%, this really messes up structural superposition for large
structures (if the structure is 300Å, this could be a 6 Å difference in
size). In my experience many groups don't seem to do this - there are quite
a lot of recent maps in the EMDB in which the nominal pixel size has been
taken as correct, and calibration with a crystal structure shows that it is
a ways off (this also affects the estimated resolution, sometimes
substantially).

Cheers,
Oli

On Mon, Feb 8, 2016 at 11:38 AM, Reza Khayat <rkhayat at ccny.cuny.edu> wrote:

> Hi Oliver,
>
>
> Out of curiosity:
>
>     1. what kind of R-factors and CC-values do you get when refining
> against the two different pixel size?
>
>     2. how different are your refined pixel sizes from one reconstruction
> to another?
>
>     ​3. how much of an affect does the wrong pixel size have on your
> downstream structure analysis (e.g. BDA, ASA, electrostatic...)?
>
>
> Best wishes,
> Reza
>
> Reza Khayat, PhD
> Assistant Professor
> City College of New York
> Department of Chemistry
> New York, NY 10031
> ------------------------------
> *From:* phenixbb-bounces at phenix-online.org <
> phenixbb-bounces at phenix-online.org> on behalf of Oliver Clarke <
> olibclarke at gmail.com>
> *Sent:* Monday, February 8, 2016 4:28 AM
> *To:* phenixbb at phenix-online.org
> *Subject:* [phenixbb] Refine pixel size of map (for EM data)?
>
> Hello,
>
> I wonder whether it would be possible to add an option for
> phenix.real_space_refine to allow refinement of the pixel size of the map
> (or the unit cell dimensions - just an overall size scale factor), and
> write out the altered map at the end of refinement.
>
> Although we try to calibrate this as best as we are able at the time of
> data collection, it is never perfect - for example, in one case I have
> dealt with, our nominal pixel size out of the scope is 1.19 Å, but the
> pixel size calibrated based on a crystal structure of a fragment of the
> protein is 1.25 Å. This is not a huge difference, but it is sufficient I
> think to have a substantial impact on refinement, particularly as regards
> clash assessment and H-bond/sec struc restraints.
>
> In cases where one does not have a solved crystal structure to use for
> calibration, perhaps refining the pixel size in conjunction with the
> geometry might be of some use?
>
> Cheers,
> Oli
>
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