[3dem] Resolution estimate: FSC vs PDB modeling

[Qiu-Xing Jiang]

Also the monomer crystal structure we determined was in solution. Our
filament formation was a membrane-dependent process, and we detected lipid
association in the isolated filaments by TLC. These complicated PDB
modeling and could not be easily ignored in the checking.

Qiu-Xing


On 3/12/14 3:01 PM, "Penczek, Pawel A" <Pawel.A.Penczek at uth.tmc.edu> wrote:

>Resolution discussion again?
>Please count me in!
>
>Regards,
>Pawel Penczek
>
>On Mar 12, 2014, at 12:59 PM, "Edward Egelman"
><egelman at virginia.edu<mailto:egelman at virginia.edu>> wrote:
>
>I feel compelled to respond to this, as I am the "senior colleague". The
>paper under discussion appeared recently in Nature (2014), Mukherjee et
>al. Let me correct the record. If one compares the map (EMDB-5795) with
>the atomic model (4MTH) all correlation disappears at better than about
>16 Angstroms, so the statement that there was strong correlation at 9.2
>to 15 Angstroms is wrong. I urge others to make this comparison
>themselves. This led me to raise questions about the 9.2 Angstrom
>resolution claim for the map. As far as I can see, after more than 20
>emails exchanged with Dr. Jiang before the submission of the paper, this
>resolution claim was simply fraudulent. That was why I asked for my name
>to be removed from this paper! I would welcome, however, for the cryo-EM
>field to move to a "reality-based" standard of resolution, and this paper
>provides a good basis for discussing why such a standard is needed.
>Regards,
>Ed Egelman
>
>On 3/12/14, 8:47 AM, Qiu-Xing Jiang wrote:
>Dear colleagues,
>This has been a topic discussed in a couple of map validation papers. I
>have a scenario encountered in a recent project and would like some input
>from those interested.
>We had a filament complex reconstruction, made of a small protein whose
>crystal structure contains mainly loops held together in the core by
>three pairs of disulfide bonds. At the time of resolution estimation, we
>used two independent maps calculated from either randomly selected halves
>or top/bottom halves to calculate FSC as usual in SPIDER. The FSC0.5 of
>the former gives 9.2 angstroms, which is more conservative than the
>FSC0.143 of the latter, and was used as a nominal estimate.
>
>The other opinion  from a senior colleague was PDB modeling, whose
>operations are to dock the X-ray structures of individual units into the
>map (assuming no change), filter the resulted PDB model to different
>resolutions, and visually determine at which resolution the map
>calculated from the PDB model and the experimental density map match the
>best(at certain thresholds), and could then be used for resolution
>estimate. When we were operating this procedure, we knew that our
>filaments had lipids associated and our map was not good enough to
>resolve the loops on the surface of the individual units in the filament.
>To make  the two maps match well to our eyes, we had to filter both to
>12-15 angstroms, which would then say the resolution was 12-15 A. The
>calculated cross-correlation between the map from the pdb model and the
>experimental map was high at 9.2-15 range, but we were not sure whether
>it would be decisively meaningful.
>
>We had debates and disagreements on which to report. At the end we
>decided to use FSC0.5 as usual, and refrained from interpreting the map
>with mutations at the subunit interface due to the experience of PDB
>modeling. I  wonder if some of you had similar experience, and more
>generally whether PDB modeling is suitable to replace FSC.
>
>Thanks for sharing your experience.
>
>Best regards,
>
>Qiu-Xing
>
>
>
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