Column 'aminoacid' error importing a Skyline ZIP file into Panorama

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Column 'aminoacid' error importing a Skyline ZIP file into Panorama Nathan Manes  2021-06-28 17:29:17
 
Hi.

The ZIP:
NIH_NitaLazar / Manes_BMDM_TLR-Chemotaxis_2021 / Manes_BMDM_TLR-Chemotaxis_2021-05-31.sky.zip

Uploading the Skyline "Share" ZIP worked fine. When I clicked "Import Data", It failed. Snippet of the log file:

28 Jun 2021 17:02:25,214 INFO : Expanding Skyline_2019-12-05_001.sky
28 Jun 2021 17:02:25,353 INFO : 1% Done
28 Jun 2021 17:03:03,307 ERROR: Import failed
28 Jun 2021 17:03:03,355 INFO : Failed to complete task 'org.labkey.targetedms.pipeline.TargetedMSImportTask'
28 Jun 2021 17:03:03,357 ERROR: aminoacid: Value is too long for column 'aminoacid', a maximum length of 30 is allowed. Supplied value was 64 characters long.

Thanks,
Nathan
 
 
vsharma responded:  2021-06-28 23:32:05
Hi Nathan,

Thank you for pointing me to your Skyline document. You have defined two neutral-loss-only modifications (Water Neutral Loss and Ammonia Neutral Loss) in your document and allowed them on all amino acids. This is exceeding the 30 characters limit that we currently have on the Panorama database column that stores the amino acid specificity for a modification. We will increase the size limit on the column so that we can support modifications that may be applied to all amino acids.

Skyline has built-in definitions for ammonia and water loss that you can select from the drop-down list in the "Edit Structural Modification" form:
Ammonia Loss (K, N, Q, R)
Water Loss (D, E, S, T)
I believe the consensus is that a fragment can exhibit ammonia loss (-NH3) if it contains the residues K, R, Q or N. And, a fragment can exhibit water loss (-H2O) if it contains the residues S,T, E or D. Is there a reason that your modification definitions allow all amino acids? If you are able to use the built-in definitions instead then you will be able to import your document into Panorama even before we make the database update.

One more thing: your .sky.zip file is 27.9GB. You can reduce the size significantly by removing unused chromatograms from your document. In Skyline go to Edit > Manage Results. Click the "Minimize" button and check the "Discard unused chromatograms" box and save. This should reduce the size of your .skyd (Skyline cache) file from ~29GB to ~416MB.

Thanks,
Vagisha
 
Nathan Manes responded:  2021-06-29 06:21:23
Hi.

Thanks for your quick reply. I didn't know about Skyline's built-in definitions for ammonia and water losses. I'll use them from now on. Since I analyzed purified synthetic peptides, I'm confident in the quality of the spectral libraries, but these definitions can only improve the data.

I did consider shrinking the SKYD, but I didn't realize that it would shrink that much, so I didn't bother.

Thanks again,
Nathan
 
Brendan MacLean responded:  2021-06-29 07:51:20

Hi Nathan,
We don't recommend non-specific ammonia and water loss definitions, and as Vagisha notes, our definition (provided by Jimmy Eng) is the closest thing to the standard in the field.

https://pubs.acs.org/doi/pdf/10.1021/pr070479v

Matrix Science:
http://www.matrixscience.com/help/fragmentation_help.html

MaxQuant:
https://pubs.acs.org/doi/pdf/10.1021/pr101065j

Jimmy also reported "ProteinProspector uses KQR for ammonia and ST for water loss amino acids"

We have in the past used non-specific losses to deal with spectral libraries from Spectronaut, but recently worked with the Biognosys team to understand exactly how they were defining these losses, which lead to the realization that they were using HQRN for ammonia loss ST for water loss. The subset definitions like KQR and ST are not a problem for Skyline, but the HQRN adding H was confusing a lot of people. The end result is that Biognosys agreed to switch to more representative KQRN probably now out in the latest version of Spectronaut.

That may be more than you wanted to know, but I thought it would be good to add this detail here where others can find it.

The short answer then is:

  1. Use the Skyline provided definitions
  2. Unless you are importing an older Spectronaut library and then you should use a slightly modified definition with K replaced with H "Ammonia Loss Spectronaut (H, Q, R, N)"

Good luck with your research. Thanks for posting this issue to the PanoramaWeb support board.

--Brendan