Searches keep failing because "An error occurred: Two modifications found site 0 of peptide". Is there any way to fix this?

[PVlasov93]

I'm trying to run a search but the following error message keeps coming up a the end. I have no idea if this can be fixed with anything on my end.
The last messages in the search screen are:
Sat Aug 23 21:11:56 EDT 2025 Importing Data Canceled!
Sat Aug 23 21:11:56 EDT 2025 An error occurred: Two modifications found (Pyrolidone from carbamidomethylated C and Acetylation of K) at site 0 of peptide CKEMVTVRSGK.
Sat Aug 23 21:11:56 EDT 2025 Please contact the developers (https://github.com/compomics/peptide-shaker/issues).

The "Acetylation of K" is a variable modification that I'm including in my SearchGui list, but I have no idea why "Pyrolidone from carbamidomethylated C" is there. The modifications I used last time were: Carbamidomethylation of C (fixed), acetylation of K, acetylation of protein N-terminus, oxidation of M, oxidation of P, citrullination of R.

If there is some issue with a single peptide in the search, is there a way to make the program omit it instead of crashing?

[hbarsnes]

Can you make sure that you are using the most recent release of PeptideShaker (v3.0.12)? If not, please update and see if that helps.

The pyrolidone modification comes from the X! Tandem second pass search and is added by default. You can change this behavior via the advanced settings for X! Tandem which you access by clicking the cogwheel to the right of the X! Tandem line in the main SearchGUI window. Under "Advanced Search" you can turn on or off the modifications that X! Tandem adds by default.

Or you can of course avoid the problem by using one or more of the other search engines supported in SearchGUI instead.

[PVlasov93]

I forgot to mention this, but I've been using four engines in my SearchGUI experiments so far: X!Tandem, MSGF+, OMSSA, and Comet. This was because another study I was citing did the same. I had previously tried removing other engines' results from the Peptide Shaker analysis, but I didn't know X!Tandem was the cause. I will try running without it.

As for the version, I recently ran the "mamba update" command for both SearchGUI and Peptide Shaker, but I will double-check that the version I got via Bioconda is the most recent.