FLASHDeconv performs ultrafast deconvolution of top down proteomics MS datasets. FLASHDeconv takes mzML file as input and outputs deconvolved feature list (.tsv) and deconvolved spectra files (.tsv, .mzML, .msalign, .ms1ft). FLASHDeconv uses SpectralDeconvolution for spectral level deconvolution and MassFeatureTracer to detect mass features. Also for MSn spectra, the precursor masses (not peak m/zs) should be determined and assigned in most cases. This assignment can be done by tracking MSn-1 spectra deconvolution information. Thus FLASHDeconv class keeps MSn-1 spectra deconvolution information for a certain period for precursor mass assignment in DeconvolvedSpectrum class. In case of FLASHIda runs, this precursor mass assignment is done by FLASHIda. Thus FLASHDeconv class simply parses the log file from FLASHIda runs and pass the parsed information to DeconvolvedSpectrum class.

See https://openms.de/FLASHDeconv for more information.

The command line parameters of this tool are:

FLASHDeconv -- Ultra-fast high-quality deconvolution enables online processing of top-down MS data
Full documentation: http://www.openms.de/doxygen/nightly/html/TOPP_FLASHDeconv.html
Version: 3.1.0-pre-feature-FLASHDeconvDevelop-2023-12-05 Dec  5 2023, 14:17:23, Revision: da80f56
To cite OpenMS:
 + Rost HL, Sachsenberg T, Aiche S, Bielow C et al.. OpenMS: a flexible open-source software platform for 
   mass spectrometry data analysis. Nat Meth. 2016; 13, 9: 741-748. doi:10.1038/nmeth.3959.
To cite FLASHDeconv:
 + Jeong K, Kim J, Gaikwad M et al.. FLASHDeconv: Ultrafast, High-Quality Feature Deconvolution for Top-Down 
   Proteomics. Cell Syst 2020 Feb 26;10(2):213-218.e6. doi:10.1016/j.cels.2020.01.003.

Usage:
  FLASHDeconv <options>

This tool has algorithm parameters that are not shown here! Please check the ini file for a detailed descript
ion or use the --helphelp option

Options (mandatory options marked with '*'):
  -in <file>*                 Input file (mzML) (valid formats: 'mzML')
  -out <file>*                Default output tsv file containing deconvolved features (valid formats: 'tsv')
  -out_spec1 <file>           Output tsv file containing deconvolved MS1 spectra. Likewise, use -out_spec2, 
                              ..., -out_spec4 to specify tsv files for MS2, ..., MS4. (valid formats: 'tsv')
  -out_mzml <file>            Output mzml file containing deconvolved spectra (of all MS levels) (valid forma
                              ts: 'mzML')
  -out_quant <file>           Output tsv file containing isobaric quantification results for MS2 only (valid 
                              formats: 'tsv')
  -out_annotated_mzml <file>  Output mzml file containing annotated spectra. For each annotated peak, monoiso
                              topic mass, charge, and isotope index are stored as meta data. Unannotated peak
                              s are also copied as well without meta data. (valid formats: 'mzML')
  -out_msalign1 <file>        Output msalign (topFD and ProMex compatible) file containing MS1 deconvolved 
                              spectra. Likewise, use -out_msalign2 for MS2 spectra. The file names for MS1 
                              and MS2 should end with ms1.msalign and ms2.msalgin respectively to be able to 
                              be recognized by TopPIC GUI.  (valid formats: 'msalign')
  -out_feature1 <file>        Output feature (topFD compatible) file containing MS1 deconvolved features. 
                              Likewise, use -out_feature2 for MS2 features. The MS1 and MS2 feature files 
                              are necessary for TopPIC feature intensity output. (valid formats: 'feature')
  -keep_empty_out             If set, empty output files (e.g., *.tsv file when no feature was generated) 
                              are kept.
  -write_detail               To write peak information per deconvolved mass in detail or not in tsv files 
                              for deconvolved spectra. If set to 1, all peak information (m/z, intensity, 
                              charge and isotope index) per mass is reported.
                              
Common TOPP options:
  -ini <file>                 Use the given TOPP INI file
  -threads <n>                Sets the number of threads allowed to be used by the TOPP tool (default: '1')
  -write_ini <file>           Writes the default configuration file
  --help                      Shows options
  --helphelp                  Shows all options (including advanced)

The following configuration subsections are valid:
 - FD       FLASHDeconv algorithm parameters
 - SD       Spectral deconvolution parameters
 - ft       Feature tracing parameters
 - iq       Isobaric quantification parameters
 - tagger   Tagger parameters

You can write an example INI file using the '-write_ini' option.
Documentation of subsection parameters can be found in the doxygen documentation or the INIFileEditor.
For more information, please consult the online documentation for this tool:
  - http://www.openms.de/doxygen/nightly/html/TOPP_FLASHDeconv.html

INI file documentation of this tool:

Legend:

required parameter

advanced parameter

+FLASHDeconvUltra-fast high-quality deconvolution enables online processing of top-down MS data

version3.1.0-pre-feature-FLASHDeconvDevelop-2023-12-05 Version of the tool that generated this parameters file.

++1Instance '1' section for 'FLASHDeconv'

in Input file (mzML)input file*.mzML

out Default output tsv file containing deconvolved featuresoutput file*.tsv

out_spec1 Output tsv file containing deconvolved MS1 spectra. Likewise, use -out_spec2, ..., -out_spec4 to specify tsv files for MS2, ..., MS4.output file*.tsv

out_spec2 Output tsv file containing deconvolved MS2 spectra.output file*.tsv

out_spec3 Output tsv file containing deconvolved MS3 spectra.output file*.tsv

out_spec4 Output tsv file containing deconvolved MS4 spectra.output file*.tsv

out_mzml Output mzml file containing deconvolved spectra (of all MS levels)output file*.mzML

out_quant Output tsv file containing isobaric quantification results for MS2 onlyoutput file*.tsv

out_annotated_mzml Output mzml file containing annotated spectra. For each annotated peak, monoisotopic mass, charge, and isotope index are stored as meta data. Unannotated peaks are also copied as well without meta data.output file*.mzML

out_msalign1 Output msalign (topFD and ProMex compatible) file containing MS1 deconvolved spectra. Likewise, use -out_msalign2 for MS2 spectra. The file names for MS1 and MS2 should end with ms1.msalign and ms2.msalgin respectively to be able to be recognized by TopPIC GUI. output file*.msalign

out_msalign2 Output msalign (topFD and ProMex compatible) file containing MS2 deconvolved spectra. The file name should end with ms2.msalign to be able to be recognized by TopPIC GUI. output file*.msalign

out_feature1 Output feature (topFD compatible) file containing MS1 deconvolved features. Likewise, use -out_feature2 for MS2 features. The MS1 and MS2 feature files are necessary for TopPIC feature intensity output.output file*.feature

out_feature2 Output feature (topFD compatible) file containing MS2 deconvolved features. The MS1 and MS2 feature files are necessary for TopPIC feature intensity output.output file*.feature

keep_empty_outfalse If set, empty output files (e.g., *.tsv file when no feature was generated) are kept.true, false

mzml_mass_charge0 Charge state of deconvolved masses in mzml output (specified by out_mzml)-1:1

write_detailfalse To write peak information per deconvolved mass in detail or not in tsv files for deconvolved spectra. If set to 1, all peak information (m/z, intensity, charge and isotope index) per mass is reported.true, false

min_mz-1.0 If set to positive value, minimum m/z to deconvolve.

max_mz-1.0 If set to positive value, maximum m/z to deconvolve.

min_rt-1.0 If set to positive value, minimum RT (in second) to deconvolve.

max_rt-1.0 If set to positive value, maximum RT (in second) to deconvolve.

log Name of log file (created only when specified)

debug0 Sets the debug level

threads1 Sets the number of threads allowed to be used by the TOPP tool

no_progressfalse Disables progress logging to command linetrue, false

forcefalse Overrides tool-specific checkstrue, false

testfalse Enables the test mode (needed for internal use only)true, false

+++FDFLASHDeconv algorithm parameters

ida_log log file generated by FLASHIda (IDA*.log). Only needed for coupling with FLASHIda acquisition

report_FDRfalse Report qvalues (roughly, point-wise FDR) for deconvolved masses. Decoy masses to calculate qvalues and FDR are also reported. Beta version.true, false

use_RNA_averaginefalse If set, RNA averagine model is used.true, false

preceding_MS1_count3 Specifies the number of preceding MS1 spectra for MS2 precursor determination. In TDP, the precursor peak of a MS2 spectrum may not belong to any deconvolved masses in the MS1 spectrum immediately preceding the MS2 spectrum. Increasing this parameter to N allows for the search for the deconvolved masses in the N preceding MS1 spectra from the MS2 spectrum, increasing the chance that its precursor is deconvolved.1:∞

isolation_window5.0 Default isolation window with. If the input mzML file does not contain isolation window width information, this width will be used.

forced_MS_level0 If set to an integer N, MS level of all spectra will be set to N regardless of original MS level. Useful when deconvolving datasets containing only MS2 spectra.0:∞

merging_method0 Method for spectra merging before deconvolution. 0: No merging 1: Average gaussian method to perform moving gaussian averaging of spectra per MS level. Effective to increase proteoform ID sensitivity (in particular for Q-TOF datasets). 2: Block method to perform merging of all spectra into a single one per MS level (e.g., for NativeMS datasets).0:2

+++SDSpectral deconvolution parameters

tol[10.0, 10.0] ppm tolerance for MS1, 2, ... (e.g., -tol 10.0 5.0 to specify 10.0 and 5.0 ppm for MS1 and MS2, respectively)

min_mass50.0 Minimum mass (Da)

max_mass1.0e05 Maximum mass (Da)

min_charge1 Minimum charge state for MS1 spectra (can be negative for negative mode)

max_charge100 Maximum charge state for MS1 spectra (can be negative for negative mode)

precursor_charge0 Charge state of the target precursor. All precursor charge is fixed to this value. This parameter is useful for targeted studies where MS2 spectra are generated from a fixed precursor (e.g., Native-MS). 0:∞

precursor_mz0.0 Target precursor m/z value. This option must be used with -target_precursor_charge option. Otherwise, it will be ignored. If -precursor_charge option is used but this option is not used, the precursor m/z value written in MS2 spectra will be used by default. 0.0:∞

min_cos[0.85, 0.85] Cosine similarity thresholds between avg. and observed isotope pattern for MS1, 2, ... (e.g., -min_cos 0.3 0.6 to specify 0.3 and 0.6 for MS1 and MS2, respectively)

min_snr[2.0, 2.0] SNR thresholds for MS1, 2, ... (e.g., -min_snr 1.0 0.6 to specify 1.0 and 0.6 for MS1 and MS2, respectively)

max_qvalue[1.0, 1.0] Qvalue thresholds for MS1, 2, ... Effective only when FDR estimation is active. (e.g., -max_qvalue 0.1 0.2 to specify 0.1 and 0.2 for MS1 and MS2, respectively)

allowed_isotope_error0 Allowed isotope index error for decoy and qvalue report. If it is set to 1, for example, +-1 isotope errors are not counted as false. Beta version.

+++ftFeature tracing parameters

mass_error_ppm-1.0 Feature tracing mass ppm tolerance. When negative, MS1 tolerance for mass deconvolution will be used (e.g., 16 ppm is used when -SD:tol 16).

quant_methodarea Method of quantification for mass traces. For LC data 'area' is recommended, 'median' for direct injection data. 'max_height' simply uses the most intense peak in the trace.area, median, max_height

min_sample_rate0.1 Minimum fraction of scans along the feature trace that must contain a peak. To raise feature detection sensitivity, lower this value close to 0.

min_trace_length10.0 Minimum expected length of a mass trace (in seconds). Only for MS1 (or minimum MS level in the dataset) feature tracing. For MSn, all traces are kept regardless of this value.

max_trace_length-1.0 Maximum expected length of a mass trace (in seconds). Set to a negative value to disable maximal length check during mass trace detection.

min_cos-1.0 Cosine similarity threshold between avg. and observed isotope pattern. When negative, MS1 cosine threshold for mass deconvolution will be used

+++iqIsobaric quantification parameters

typenone Isobaric Quantitation method used in the experiment.none, itraq4plex, itraq8plex, tmt10plex, tmt11plex, tmt16plex, tmt18plex, tmt6plex

isotope_correctiontrue Enable isotope correction (highly recommended). Note that you need to provide a correct isotope correction matrix otherwise the tool will fail or produce invalid results.true, false

reporter_mz_tol2.0e-03 m/z tolerance in Th from the expected position of reporter ion m/zs.

+++taggerTagger parameters

max_tag_count0 Maximum number of the tags per length (lengths set by -min_length and -max_length options). The tags with different amino acid combinations are all treated separately. E.g., TII, TIL, TLI, TLL are distinct tags even though they have the same mass differences. but are counted as four different tags. 0:∞

min_length4 Minimum length of a tag. Each mass gap contributes to a single length (even if a mass gap is represented by multiple amino acids). 3:30

max_length15 Maximum length of a tag. Each mass gap contributes to a single length (even if a mass gap is represented by multiple amino acids). 3:30

flanking_mass_tol200.0 Flanking mass tolerance in Da.

allowed_isotope_error0 Allowed_isotope_error for tag generation. It only applies to amino acids, not mass gaps.0:1

max_gap_count0 Maximum mass gap count per tag.0:2

max_aa_in_gap2 Maximum amino acid count in a mass gap.2:3

min_matched_aa6 Minimum number of amino acids in matched proteins, covered by tags.

fasta Target protein sequence database against which tags will be matched.input file

out Tagger output file.output file