iProphet failing when combining Comet and MSGF in TPP 5.2 using Ubuntu 18.04

[Alejandro]

Hello all,

I'm having trouble running TPP 5.2 in general in Ubuntu 18.04. I followed the compilation guideline, and after using svn checkout -r 7922 (as recommended by Eric)  it went ahead, although I still have some things that are broken, like checking for the ions in the pepxml file. I get the following error:

Error - scan 10000 is an MS1 scan in the mzXML file /home/laptop/Documents/TPP_data/tests/QuickYeastUPS1/UPS1_50000amol_R1.mzML

I can click on the Spectrum and see the peptide table but whenever I click on the ions I see that error.

More importantly, I can run Comet followed by PeptideProphet, iProphet and ProteinProphet and it all goes fine. I can do the same with MSGF+ (PeptideProphet, iProphet and ProteinProphet). However, when I try to combine both PeptideProphet files (from Comet and MSGF+) I always get the following error:

EXECUTING: cd /home/laptop/Documents/TPP_data/TH129_134/comsgf ; /usr/local/tpp/bin/InterProphetParser /home/laptop/Documents/TPP_data/TH129_134/Comet/interact_comet.pep.xml /home/laptop/Documents/TPP_data/TH129_134/MSGF/interact_p005.pep.xml interact.ipro.pep.xml

Running FPKM NSS NRS NSE NSI NSM NSP Model EM:
Computing NSS values ... 

Creating 1 threads 
Wait for threads to finish ...
.... done
Computing NRS values ... 

Creating 1 threads 
Wait for threads to finish ...
0--------------------------------------------------50------------------------------------------------100%
..................................................................................................... done
Computing NSE values ... 

Creating 1 threads 
Wait for threads to finish ...
0--------------------------------------------------50------------------------------------------------100%
..................................................................................................... done
Computing NSI values ... 

Creating 1 threads 
Wait for threads to finish ...
0--------------------------------------------------50------------------------------------------------100%
..................................................................................................... done
Computing NSM values ... 

Creating 1 threads 
Wait for threads to finish ...
0--------------------------------------------------50------------------------------------------------100%
..................................................................................................... done
Computing NSP values ... 
Creating 1 threads 
Wait for threads to finish ...
0--------------------------------------------------50------------------------------------------------100%
..................................................................................................... done
FPKM values are unavailable ... 
Iterations: DEBUG HERE ...
Segmentation fault (core dumped)

Command FAILED
RETURN CODE:35584 

I have tried to disable FPKM, but gives the same error. Odd is that I can run iProphet on each and runs fine, including FPKM.

My samples are dimethylated samples, and was searched using static modifications, doing one search for Light and one search for Heavy, and then both files combined with PeptideProphet., like so:

xinteract -Ninteract_comet.pep.xml -p0.05 -l7 -PPM -OAPd -dDECOY H_TH189.pep.xml L_TH189.pep.xml 

I have also tried using -p0 but gives in the end the same error when combining both search engines. 

In TPP 5.1 the combination works, however I'm trying to use the new Xpress which uses the intensity for quantitation, and in TPP 5.1 the intensities are not passed to the proteins.

Any help would be appreciated. Thanks,

Alejandro

[David Shteynberg]

Hello Alejandro,

It appears that you are coming across an iProphet error in that svn code revision.  Are you willing to test if the latest revision (7931) fails in the same way on your data?  Otherwise, if you are able to share the data I can attempt to replicate the error myself and correct the bug.

Thanks,

-David

--
You received this message because you are subscribed to the Google Groups "spctools-discuss" group.
To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.
To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.
To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/38e40045-66e2-4d7e-8acf-4a9995047464%40googlegroups.com.
For more options, visit https://groups.google.com/d/optout.

[Alejandro]

Hi David,

I installed the revision 7931 and it gives the same error:

I also tested in each file alone and I can run iProphet succesfully. However combining files from MSGF and Comet fails.

Best,

Alejandro

To unsubscribe from this group and stop receiving emails from it, send an email to spctools...@googlegroups.com.

[David Shteynberg]

Can ypu post your msgf and comet results for me to download and test.  Please let me know.  Also if you are at ASMS, stip by our booth and I can get your dataset to test there. 

Cheers,

David

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/161a30c8-2fa3-4a20-9683-c726ad147c14%40googlegroups.com.

[David Shteynberg]

Apologies for the misspellings in my last message...I am typing this on my phone

[David Shteynberg]

I was able to replicate the error on a different  dataset.  I will work on a solution in the coming days, meanwhile the older version of iProphet will work just fine.  The new version will be faster when it's released but results shouldn't change. 

Cheers,

David

On Mon, Jun 3, 2019, 4:45 AM Alejandro <agome...@gmail.com> wrote:

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/161a30c8-2fa3-4a20-9683-c726ad147c14%40googlegroups.com.

[David Shteynberg]

Hello Alejandro,

I have committed a fix for this problem.  Would you mind seeing if it works on your end?  The revision number is 7932.  Thanks for reporting the issue and helping to make the TPP better.

Cheers,

-David

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/161a30c8-2fa3-4a20-9683-c726ad147c14%40googlegroups.com.

[Alejandro]

HI David,

Sorry for the late reply, I was out of town.

Thanks for looking into it. I recompiled and installed the revision 7932. It now runs iProphet with those files! However, ProteinProphet now fails with the following error:

COMMAND 1 [MON JUN 10 18:28:57 2019]    show

EXECUTING: cd /home/laptop/Documents/TPP_data/TH189 ; /usr/local/tpp/bin/ProteinProphet /home/laptop/Documents/TPP_data/TH189/interact_comsgf_v2.ipro.pep.xml /home/laptop/Documents/TPP_data/TH189/interact_comsgf.prot.xml IPROPHET 

ProteinProphet (C++) by Insilicos LLC and LabKey Software, after the original Perl by A. Keller (TPP v5.2.1-dev Flammagenitus, Build 201906101745-7932 (Linux-x86_64))
 (no FPKM) (using iProphet probs) (using degen pep info)
Reading in /home/laptop/Documents/TPP_data/TH189/interact_comsgf_v2.ipro.pep.xml...
Error: multiple databases referenced by /home/laptop/Documents/TPP_data/TH189/interact_comsgf_v2.ipro.pep.xml
Use RefreshParser to update all input files to common database

Command FAILED
RETURN CODE:256

I have double check the Comet and MSGF pep.xml files and they map to the same database, using the same structure. In any case I remap the files, and it stills gives me the same error.

Best,

Alejandro

[David Shteynberg]

Hello Alejandro,

This is a new issue so that's good, as we have made some progress.  I am not sure why ProteinProphet is telling you you have two different databases referenced by your two searches.  Can you either post your pep.xml input file and database for me to test with ProteinProphet or post all relevant lines referencing the database from your input files?  A single file pep.xml may contain multiple MS runs and they would all have to reference the same database in the same way.  Based on my understanding of how you've run iProphet, I expect that you'd have a single input file to ProteinProphet.  Can you remap this file against the database and run ProteinProphet on that?

Thanks,

-David

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/b3b5fb8b-17e5-4c64-9964-80ca01349db6%40googlegroups.com.

[Alejandro]

Hi David,

I re-mapped it and now is working. A step forward. Now, I'm running XPRESS and I get an error when running it within iProphet, I use:

/usr/local/tpp/bin/XPressPeptideParser interact.ipro.pep.xml -m10 -a -c5 -H -b -i -nn,6.031817 -nK,6.031817

And get always:

EXECUTING: cd /home/laptop/Documents/TPP_data/TH189/comsgf ; /usr/local/tpp/bin/XPressPeptideParser interact.ipro.pep.xml -m20 -a -c5 -H -b -i -nn,6.031817 -nK,6.031817 

.................................................. 1k
.................................................. 2k
.....................Segmentation fault (core dumped)

Command FAILED
RETURN CODE:35584

I can run XPRESS successfully the same way, in either the Comet (PeptideProphet and iProphet) or the MSGF run separately. I can then run iPropeht on both PeptideProphet files and import the XPRESS file with success. However if I run XPRESS at the iProphet step when combining both files it gives that error.

Best,

Alejandro

[David Shteynberg]

Hi Alejandro,

It appears that XPress runs for awhile then crashes in the middle of the file.  This may indicate that different ms run in the pepxml file reference different mzML files to extract the data, and some of those mzML references could be incorrect.  Have you moved the files around after processing?  Can you grep your file for mzML and make sure the paths are valid?  Otherwise,  I would have to play with your data to troubleshoot the issue.

Thanks,

David 

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/156c48b8-2602-4c6e-8fd0-f6aaf9b8667c%40googlegroups.com.

[Alejandro]

Hi David,

No, I haven't moved the files. I grep mzML and it has 4 instances, one for Heavy, one for Light for Comet, as well as one Heavy and one Light for MSGF. Running each independently (Comet or MSGF+) generates no error. 

Thanks,

Alejandro

[David Shteynberg]

Hi Alejandro,

XPress might be getting confused with the way you've done the search.  Can you redo the search once, using variable heavy label modification and see if it lets you quantify the results?

Thanks,

-David

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/6c375b42-55bf-47e2-badf-7aa9ac4b3f69%40googlegroups.com.

[Alejandro]

Hi David,

The test didn't went forward not because of TPP but because of how MSGF+ encodes the modifications. I have to troubleshoot that first. However, I don't know if that will make a big change. In TPP 5.1 and below I can do XPRESS quantitation when running iProphet of both results (Comet and MSGF). I can also do it separately in TPP 5.2 : Comet > PeptideProphet > iProphet > Xpress or MSGF > PeptideProphet > iProphet > Xpress, is just when I do iProphet of the combined Comet/MSGF+ plus Xpress that it hangs. I can also run PeptideProphet with Xpress in each (Comet or MSGF) and then combine them with iProphet and I get the quantitation results in the combined iProphet file. Just I would have to run Xpress on each search engine result.

Best,

Alejandro

[David Shteynberg]

Hi Alejandro,

I am trying (unsuccessfully) to replicate this behavior on a different iProphet combination of comet and xtandem results.  Would it be possible for you to provide some test data so I can reproduce this specific bug in my environment?

Thanks,

-David

To unsubscribe from this group and stop receiving emails from it, send an email to spctools-discu...@googlegroups.com.

To post to this group, send email to spctools...@googlegroups.com.
Visit this group at https://groups.google.com/group/spctools-discuss.

To view this discussion on the web visit https://groups.google.com/d/msgid/spctools-discuss/6d7beb83-2ddf-4128-9423-f14e4fbba0a3%40googlegroups.com.