error processin mzML files

[jonathanctrinidad-blip]

We have been using 3.1 successfully to search Thermo .raw files. We now need to search mzML files (made from Thermo .raw files). However, all of those searches end up failing, even if they were made from .raw files that worked when run directly. Here is the portion of the log file detailing the error:

Any ideas what is going on? Thanks!

Process ID=06572: [pParse] - --------- BEGIN PARAMETERS -----------
Process ID=06572: [pParse] - 01: check_activationcenter = 1
Process ID=06572: [pParse] - 02: co-elute = 1
Process ID=06572: [pParse] - 03: cut_similiar_mono = 1
Process ID=06572: [pParse] - 04: datanum = 1
Process ID=06572: [pParse] - 05: datapath1 = C:\Cohen\Glycopeptide_data\pglyco_searches\F20250802-08.mzML
Process ID=06572: [pParse] - 06: delete_msn = 0
Process ID=06572: [pParse] - 07: dia_mode = 0
Process ID=06572: [pParse] - 08: dia_mode_to_filter_by_selectedMS2ScanNoSet = 1
Process ID=06572: [pParse] - 09: input_format = mzML
Process ID=06572: [pParse] - 10: intensity = 1
Process ID=06572: [pParse] - 11: ipv_file = IPV.txt
Process ID=06572: [pParse] - 12: isolation_width = 2.000000
Process ID=06572: [pParse] - 13: logfilepath = C:\Cohen\Glycopeptide_data\pglyco_searches
Process ID=06572: [pParse] - 14: m/z = 5
Process ID=06572: [pParse] - 15: mars_model = 4
Process ID=06572: [pParse] - 16: mars_threshold = -0.500000
Process ID=06572: [pParse] - 17: mstol = 20.000000
Process ID=06572: [pParse] - 18: mstolppm = 1
Process ID=06572: [pParse] - 19: output_mgf = 1
Process ID=06572: [pParse] - 20: output_pf = 1
Process ID=06572: [pParse] - 21: outputpath =
Process ID=06572: [pParse] - 22: recalibrate_window = 7.000000
Process ID=06572: [pParse] - 23: rewrite_files = 0
Process ID=06572: [pParse] - --------- END PARAMETERS -------------
Process ID=06572: [pParse] - .\xtract_xml.exe -ms -a -i 1 -m 5 -o "C:\Cohen\Glycopeptide_data\pglyco_searches" "C:\Cohen\Glycopeptide_data\pglyco_searches\F20250802-08.mzML"
Process ID=06572: '.\xtract_xml.exe' is not recognized as an internal or external command,
Process ID=06572: operable program or batch file.
Process ID=06572: [pParse] - Cannot use xtract to extract signals from: C:\Cohen\Glycopeptide_data\pglyco_searches\F20250802-08.mzML
Process ID=06572: [pParse] - pParse has encountered a problem and will exit.
Process ID=06572: You can send your pParsePlusLog.txt to pparse@ict.ac.cn for help.
Process ID=06572: [pGlyco] Processing 1 raw files.
Process ID=06572: C:\pGlyco3.1>pGlycoDB.exe "C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlyco.cfg"
Process ID=06572: *****************************************************
Process ID=06572: * *
Process ID=06572: * * Welcome to pGlyco3 * *
Process ID=06572: * *
Process ID=06572: * * pglyco@ict.ac.cn * *
Process ID=06572: * *
Process ID=06572: *****************************************************
Process ID=06572: [pGlyco] pGlyco software will be expired in 2027/01/01
Process ID=06572: [pGlyco] top_n_peaks=300
Process ID=06572: [pGlyco] output_top_n=5
Process ID=06572: [pGlyco] --. [key-config]
Process ID=06572: [pGlyco] 1. aaini=aa.ini
Process ID=06572: [pGlyco] 2. glycoini=glyco.ini
Process ID=06572: [pGlyco] 3. modini=modification.ini
Process ID=06572: [pGlyco] --. [glycan]
Process ID=06572: [pGlyco] 4. glycan_type=N-glycan
Process ID=06572: [pGlyco] 5. glycosylation_sites=J
Process ID=06572: [pGlyco] 6. glycan_core=N(1),N(2),N(2)H(1),N(2)H(2),N(2)H(3),N(1)F(1),N(2)F(1)
Process ID=06572: [pGlyco] 7. glycan_db=pGlyco-N-Human.gdb
Process ID=06572: [pGlyco] 8. glycan_fix_mod=
Process ID=06572: [pGlyco] 9. glycan_var_mod=
Process ID=06572: [pGlyco] 10. max_var_mod_on_glycan=1
Process ID=06572: [pGlyco] 11. max_glycan_db_size=100000
Process ID=06572: [pGlyco] 12. gp_marker_glyco=N
Process ID=06572: [pGlyco] --. [protein]
Process ID=06572: [pGlyco] 13. fasta=C:\Users\msfuser\Desktop\Homo_sapiens_proteome_2024.fasta.N2J
Process ID=06572: [pGlyco] 14. enzyme=Trypsin Nterm _ _ Cterm KR _
Process ID=06572: [pGlyco] 15. max_miss_cleave=3
Process ID=06572: [pGlyco] 16. protein_fix_mod=Carbamidomethyl[C]
Process ID=06572: [pGlyco] 17. protein_var_mod=Oxidation[M],Acetyl[ProteinN-term],Gln->pyro-Glu[AnyN-termQ],Met-loss+Acetyl[ProteinN-termM],Met-loss[ProteinN-termM]
Process ID=06572: [pGlyco] 18. max_var_mod_on_peptide=2
Process ID=06572: [pGlyco] --. [search]
Process ID=06572: [pGlyco] 19. fragmentation_type=HCD
Process ID=06572: [pGlyco] 20. precursor_tolerance=10 ppm
Process ID=06572: [pGlyco] 21. fragment_tolerance=20 ppm
Process ID=06572: [pGlyco] 22. output_dir=C:/Cohen/Glycopeptide_data/pglyco_searches/802-08-mzml_test/process1/
Process ID=06572: [pGlyco] Loading glycan database ...
Process ID=06572: [pGlyco] 2920 glycan structures
Process ID=06572: [pGlyco] Ion Indexing ...
Process ID=06572: [pGlyco] Ion Indexing done
Process ID=06572: [pGlyco] Glycan grouping ...
Process ID=06572: [pGlyco] Glycan grouping done.
Process ID=06572: [pGlyco] Loading protein sequences ...
Process ID=06572: [pGlyco] 590776 peptide sequences ...
Process ID=06572: [pGlyco] Preparing spectra ...
Process ID=06572: [pGlyco Error] Open in CMGFReader: Could not open file: C:\Cohen\Glycopeptide_data\pglyco_searches\F20250802-08_HCDFT.mgf
Process ID=06572: C:\pGlyco3.1>gpPercolator.exe -p "C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlyco.cfg"
Process ID=06572: C:\pGlyco3.1>pGlycoFDR.exe -p "C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlyco.cfg"
Process ID=06572: C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlycoDB-GP-Raw1-FDR.txt
Process ID=06572: [pGlycoFDR] 0 GPSMs at 10.0% FDR
Process ID=06572: C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlycoDB-GP-Raw1-FDR-noFiltered.txt
Process ID=06572: [pGlycoFDR] 0 GPSMs at 100.0% FDR
Process ID=06572: merge into C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlycoDB-GP-FDR.txt
Process ID=06572: C:\pGlyco3.1>pGlycoProInfer.exe -p "C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlyco.cfg"
Process ID=06572: Reading pGlyco results ...
Process ID=06572: Inferring proteins ...
Process ID=06572: C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlycoDB-GP-FDR-Pro.txt
Process ID=06572: C:\pGlyco3.1>XIC.exe -p "C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlyco.cfg"
Process ID=06572: [XIC] Smoothing window = 21
Process ID=06572: [XIC] Smoothing method = savgol_filter
Process ID=06572: [XIC] Loading pGlyco results ...
Process ID=06572: [XIC] RT window is [-120.0, +120.0] seconds
Process ID=06572: [XIC] ms1 file of raw "c:\cohen\glycopeptide_data\pglyco_searches\f20250802-08" not found
Process ID=06572: C:\pGlyco3.1>pGlycoSite.exe "C:\Cohen\Glycopeptide_data\pglyco_searches\802-08-mzml_test\process1\pGlyco.cfg"
Process ID=06572: Already registered!
Process ID=06572: [pGlycoSite] Glycosylation site localization finished!
[pGlyco] All results are merged!
[pGlyco] Running time = 0.2 minutes.

[jonathanctrinidad-blip]

To note: we have tried making the mzML files in both Proteome Discoverer and MSConvert. Neither version worked on pGlyco