Error from importing Peptide Shaker mzid lauh2  2021-03-30
 

Hello Skyline team,

I am trying to build a library using the mzid from Peptide Shaker. However, Skyline (64-bit) 20.2.0.343 (a7a9e8c4f) is giving me the following error,

System.IO.IOException: ERROR: No spectra were found for the new library.

Command-line: C:\Users\HoTak.Lau\AppData\Local\Apps\2.0\8MPRNX2K.0X7\9EA9LC5J.M7P\skyl..tion_e4141a2a22107248_0014.0002_2f1cb11a037aa924\BlibBuild -s -A -H -o -c 0.99 -i testShakerOut -S "C:\Users\HoTak.Lau\AppData\Local\Temp\tmp4C0F.tmp" "C:\Users\HoTak.Lau\Desktop\MS_tools\PeptideShaker-2.0.16\resources\example_dataset\data\testShakerOut.redundant.blib"
Working directory: C:\Users\HoTak.Lau\Desktop\MS_tools\PeptideShaker-2.0.16\resources\example_dataset\data
   at pwiz.Common.SystemUtil.ProcessRunner.Run(ProcessStartInfo psi, String stdin, IProgressMonitor progress, IProgressStatus& status, TextWriter writer) in C:\proj\skyline_20_2_x64\pwiz_tools\Shared\Common\SystemUtil\ProcessRunner.cs:line 62
   at pwiz.BiblioSpec.BlibBuild.BuildLibrary(LibraryBuildAction libraryBuildAction, IProgressMonitor progressMonitor, IProgressStatus& status, String& commandArgs, String& messageLog, String[]& ambiguous) in C:\proj\skyline_20_2_x64\pwiz_tools\Shared\BiblioSpec\BlibBuild.cs:line 201
   at pwiz.Skyline.Model.Lib.BiblioSpecLiteBuilder.BuildLibrary(IProgressMonitor progress) in C:\proj\skyline_20_2_x64\pwiz_tools\Skyline\Model\Lib\BiblioSpecLiteBuilder.cs:line 156

I generate the mzid using PeptideShaker 2.0.16 (2.0.14 and 2.0.18 gave the same error) -> Open Example -> Export -> Follow Up Analysis -> Skyline Export -> Export as mzid, to the PeptideShaker example folder, as mzid v1.1.gzip -> extract the .mzid using 7zip -> import to Skyline.

Is there anyway to get around this?

Thank you
Ho-Tak

 
 
Kaipo responded:  2021-03-30

Hi Ho-Tak,
This may mean that the cut-off score used for building the library was too restrictive. Would you mind attaching the mzid file to this request (or uploading it to https://skyline.ms/files.url if it is large)?

Thanks,
Kaipo

 
Brendan MacLean responded:  2021-03-30

Since the reported command-line is:

BlibBuild -s -A -H -o -c 0.99 -i testShakerOut -S "C:\Users\HoTak.Lau\AppData\Local\Temp\tmp4C0F.tmp" "C:\Users\HoTak.Lau\Desktop\MS_tools\PeptideShaker-2.0.16\resources\example_dataset\data\testShakerOut.redundant.blib"

The cut-off score is 0.99, which seems okay. I am a bit curious why the input file appears to be "C:\Users\HoTak.Lau\AppData\Local\Temp\tmp4C0F.tmp"

Is it possible you are still pointing to a file which Windows believes to be a compressed ZIP file and it has extracted it to this temporary location?

Does this file path look at all familiar to you?

 
lauh2 responded:  2021-03-30

hmmmm. I am not sure, I moved all of the files to a new folder in Desktop; changed the Cut-off score to 0.5, and tried again. But it is still giving me "the same" error.

System.IO.IOException: ERROR: No spectra were found for the new library.

Command-line: C:\Users\HoTak.Lau\AppData\Local\Apps\2.0\8MPRNX2K.0X7\9EA9LC5J.M7P\skyl..tion_e4141a2a22107248_0014.0002_2f1cb11a037aa924\BlibBuild -s -A -H -o -c 0.5 -i desktopFolder -S "C:\Users\HoTak.Lau\AppData\Local\Temp\tmpBE03.tmp" "C:\Users\HoTak.Lau\Desktop\testDesktop\desktopFolder.redundant.blib"
Working directory: C:\Users\HoTak.Lau\Desktop\testDesktop
   at pwiz.Common.SystemUtil.ProcessRunner.Run(ProcessStartInfo psi, String stdin, IProgressMonitor progress, IProgressStatus& status, TextWriter writer) in C:\proj\skyline_20_2_x64\pwiz_tools\Shared\Common\SystemUtil\ProcessRunner.cs:line 62
   at pwiz.BiblioSpec.BlibBuild.BuildLibrary(LibraryBuildAction libraryBuildAction, IProgressMonitor progressMonitor, IProgressStatus& status, String& commandArgs, String& messageLog, String[]& ambiguous) in C:\proj\skyline_20_2_x64\pwiz_tools\Shared\BiblioSpec\BlibBuild.cs:line 201
   at pwiz.Skyline.Model.Lib.BiblioSpecLiteBuilder.BuildLibrary(IProgressMonitor progress) in C:\proj\skyline_20_2_x64\pwiz_tools\Skyline\Model\Lib\BiblioSpecLiteBuilder.cs:line 156

The .7z attached here has the .mzid and .mzML from Peptide Shaker and the Skyline files created from trying to import the .mzid.

 
lauh2 responded:  2021-03-30

Sorry. The .7z is too large. I uploaded to the file sharing page.

 
Kaipo responded:  2021-03-30

Hi Ho-Tak,
Skyline is having trouble matching spectra in the mzid to the corresponding ones in the mzml file. For PeptideShaker mzid files, we use the spectrumID attribute of the SpectrumIdentificationResult to match spectra, but it doesn't seem to be useful in this case.
I did notice that the PSMs in the mzid have "spectrum title" which match spectra in the mzml, so I will make a fix for the next release to use these values. Until then, you may be able to workaround the issue by replacing the spectrumID values with the "spectrum title" values (via scripting/programatically, it is probably not feasible manually).

Thanks,
Kaipo

 
lauh2 responded:  2021-03-30

Hi Kaipo,

Thank you for looking into it. I will wait for the next release.

Ho-Tak

 
matt.chambers42 responded:  2021-03-31

Hi Ho-Tak,

What was the input to PeptideShaker? My understanding is PS only supports MGF spectrum files. Did you have an MGF intermediate file as well as mzML? Perhaps BiblioSpec is preferring the mzML even though the MGF is there? In that case we should change the PS mzIdentML reader to prefer MGF (or perhaps ONLY support MGF, like PS itself).

-Matt

 
Kaipo responded:  2021-03-31

Also related, there is actually a way you can build the library without any bug fixes.
msconvert (part of the ProteoWizard project: http://proteowizard.sourceforge.net/download.html) can be used to convert the mzML file to MGF format.
Then, replace the mzML file with the converted MGF and the library should build successfully.

 
lauh2 responded:  2021-04-01

Hi Skyline Team,

Sorry for the late reply.
I tested the whole workflow again. And it is working now.

To get it to work, I have to convert a Thermo RAW using MSconvert (peak picking) to .mgf -> do peptide search using SearchGUI, and build a Peptide Shaker project -> export .mzID for Skyline -> import to Skyline.

I tried to start with an mzML (converted from Thermo RAW), peptide search, and convert .mzML to .mgf. This way doesn't work.

So I think we can close this case now.

Thank you.

 
matt.chambers42 responded:  2021-04-01

Hmm, so you're saying this works:

Thermo RAW -> msconvert -> MGF -> SearchGUI -> PeptideShaker -> mzIdentML -> Skyline
                            \--------------------------------------------------/

But this doesn't:

Thermo RAW -> msconvert -> mzML -> SearchGUI -> PeptideShaker -> mzIdentML -> Skyline
                             \----------> msconvert -> MGF ---------------------/

For the top route, is an mzML file created at all?

 
lauh2 responded:  2021-04-01

Correct. No mzML was created at all.

Interestingly, if I convert the example mzML shipped by Peptide Shaker (the one I uploaded) to mgf, then it will work. But not my own RAW -> mzML -> MGF.