[3dem] Resolution estimate: FSC vs PDB modeling

[Penczek, Pawel A]

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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