Error in PeptideProphet via FragPipe when using custom enzyme

[jossmi]

I am using MSFragger 3.1.1, Philopher 3.4.13, and FragPipe 14.0 in Windows.

I am doing an open search and have digested my samples using a fairly uncommon enzyme (wild-type alpha-lytic protease, aka WALP; citation: 10.1074/mcp.M113.034710), which cleaves after TASV. In the MsFragger tab of FragPipe, I have selected "custom" for enzyme rules, entered "WALP" for enzyme name, and entered "TASV" for cut after. On the Validation tab of FragPipe, under PeptideProphet, i have entered: --nonparam --expectscore --decoyprobs --masswidth 1000.0 --clevel -2 --enzyme WALP : cut(TASV) sense(C)

I got the following error:

INFO[17:04:56] Executing PeptideProphet v3.4.13
terminate called after throwing an instance of 'boost::exception_detail::clone_impl<boost::exception_detail::error_info_injector<std::ios_base::failure[abi:cxx11]> >' what(): failed opening file: The filename, directory name, or volume label syntax is incorrect.​

: iostream error FATA[17:04:58] Cannot execute program. There was an error with PeptideProphet, please check your parameters and input files Process 'PeptideProphet' finished, exit code: 1 Process returned non-zero exit code, stopping

I am sure my syntax is wrong on the PeptideProphet command line, but I wanted to see if you could help. I've attached my log and parameter files.

Thanks, Josh

fragger_params.txt fragpipe_2021-01-26_17-00-13_config.txt log_2021-01-26_17-07-19.txt shepherd_config.txt umpire-se_2021-01-26_17-00-13_params.txt

[prvst]

@anesvi In this case, do you think that PeptideProphet should be set with the nonspecific enzyme type?

[anesvi]

PeptideProphet can only recognized a number of predefined enzymes. So if you create a new enzyme name in MSFragger, it will crash philosopher

So yes, you can define a new set of cleavage rules in MSFragger, but put ‘nonspecific’ in the enzyme name field. Or, alternatively, add --enzyme nonspecific to PeptideProphet command in Validation tab. I think peptideprophet will run fine, and you do not need to rerun MSFragger. Re-Start from Validation

Alexey

From: Felipe Leprevost notifications@github.com Sent: Wednesday, January 27, 2021 9:50 AM To: Nesvilab/philosopher philosopher@noreply.github.com Cc: Nesvizhskii, Alexey nesvi@med.umich.edu; Mention mention@noreply.github.com Subject: Re: [Nesvilab/philosopher] Error in PeptideProphet via FragPipe when using custom enzyme (#180)

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@anesvihttps://github.com/anesvi In this case, do you think that PeptideProphet should be set with the nonspecific enzyme type?

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[jossmi]

Alexey and Felipe, thanks a ton!

[jossmi]

Hi, sorry to add another comment after this was closed - the "nonspecific" enzyme name in MSFragger and omitting any "--enzyme" note in PeptideProphet on the validation tab worked to get me through PrptideProphet. However, now I am getting an error with PTMShepherd:

Creating combined histogram 11:29:01.711 [main] WARN edu.umich.andykong.ptmshepherd.peakpicker.PeakSummary - Empty 'features' encountered Exception in thread "main" java.lang.NullPointerException at edu.umich.andykong.ptmshepherd.peakpicker.ModSummary.toFile(ModSummary.java:104) at edu.umich.andykong.ptmshepherd.PTMShepherd.main(PTMShepherd.java:379) Process 'PTMShepherd' finished, exit code: 1 Process returned non-zero exit code, stopping Generated histogram file for dataset default-ptmshepherd-dataset [-2147483648 - -2147483648]

Any idea what's going on? I've attached my log file. With the exception of the enzyme settings, all the parameters are the same as what I attached before.

log_2021-01-27_11-31-09.txt

Thanks, Josh

[prvst]

Hi @jossmi. Feel free to keep posting if necessary. I'll re-open the issue and move it to the FragPipe repository so others (including the person responsible for PTM-Shepherd) can help you too.

[jossmi]

Thanks, Felipe.

Just to be a bit more clear on my enzyme settings, In MSFragger, I selected "custom" from the dropdown menu, entered "nonspecific" in the enzyme name. See screenshot.

Josh

[fcyu]

This issue has been solved. There is a new issue created.

Best,

Fengchao

[jossmi]

Fengchao, sorry, I'm assuming you're referring to a new issue I posted in in PTM-S about a similar, but separate PTM-S issue (that was with a regular Trypsin digestion, this is with an uncommon enzyme; I'm unfortunately juggling a couple different issues). The issue for this particular thread has not been resolved yet.

I still would like any input on how to get PTM-S to accept my custom digestion rules (cut after TASV).

Thanks, Josh

[fcyu]

Hi Josh,

Sorry for the oversight. I will re-open it.

Best,

Fengchao

[danielgeiszler]

PTM-Shepherd does not do anything with enzymes. This is an unrelated issue to the one you opened the ticket for.

You only have 359 PSMs. This is simply not enough data to run PTM-Shepherd when you have such a large mass range. If you are certain your parameters are correct, I would suggest disabling it.

We can continue discussing in the other thread, because it is the same issue as this one.