Mascot .dat file failed to run peptide and protein prophet

[Vidya Venkatraman]

Hi,

I have encountered an issue wherein if I try to convert a mascot .dat file to pepxml, it does the conversion and when I try to use this pepxml  to run peptideprophet and proteinprophet, it fails. 

I tried to check if .dat to pepxml conversion worked properly, but realised that when I click on the pepxml, nothing loads on the pepxml viewer and I also saw that usually there is an index file that is created and in this case, it did not create the index file as well. Just to make sure the .dat file is good, i was also able to open it up on Scaffold and see some proteins.

This is the link to the shared dropbox folder called "TPP"

https://www.dropbox.com/sh/4unsley0gir7jve/7D3-XANnj8

I have 2 folders there, one called "working files" where I tested a .dat file (from same samle, different fraction) conversion to pepxml and ran peptide and protein prophet and it worked successfully. The other folder called "not working files" has the .dat file which failed to run peptide and protein prophet. Also attached are .mzxml, .pepxml, the log of conversion from .dat to .pepxml and errorlog of conversion from pepxml to peptide and protein prophet along with screenshots. I have also included the fasta file I used in the "Fasta" folder. 

Can you anyone guide me as to what could have gone wrong?

Thanks in advance,

Vidya

[Joseph Slagel]

Hi Vidya,

Thanks for providing such great detail on the issue and I'm sorry it isn't working for you.  We're taking a look at what you posted and there is an error indicating invalid XML formatting in one of your files.  Unfortunately this is one of the ones that you haven't posted to dropbox.  Any chance you can upload the following file from your system (from the "not working" case)?

C:/Inetpub/wwwroot/ISB/data/data/interact.pep.xml"

Thanks,
Joe

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[Vidya Venkatraman]

Hi Joe,

Thanks for your quick response. I have put all the remaining output files into dropbox now. Please take a look and let me know if you need any other information.

Vidya

[Joseph Slagel]

Vidya,

I noticed that you happen to have the string "<alpha>" in some of your sequence definition lines in your database.  These are showing up in your interact.pep.xml and causing the parser to fail.  Now we should be escaping anything that looks like xml in the sequence definition its possible we missed it somewhere.  Could you try just removing the "<>" from the descriptions in the fasta file and rerunning?  The four lines are:

>tr|Q9UMA4|Q9UMA4_HUMAN IFN-<alpha>2 protein (Fragment) OS=Homo sapiens GN=IFN-<alpha>2 PE=2 SV=1
>tr|A0N7E2|A0N7E2_HUMAN V<alpha>13 protein (Fragment) OS=Homo sapiens GN=V<alpha>13 PE=2 SV=1
>tr|A0N7E4|A0N7E4_HUMAN V<alpha>1 protein (Fragment) OS=Homo sapiens GN=V<alpha>1 PE=2 SV=1

-Joe

To view this discussion on the web visit https://groups.google.com/d/msg/spctools-discuss/-/sR6TGOvkegwJ.

[Vidya Venkatraman]

HI Joe,

That seems to have resolved the problem. The issue seems to have been there in the pepxml itself so I went back and redit the .dat to pepxml conversion with the edited fasta file and now I saw that the pepxml viewer actually loaded. Using this pepxml file, the peptide and protein prophet works.

If I run into any further issues, I will post back.

Thanks so much,
Vidya