PeptideSpectrumMatching

PeptideSpectrumMatching identifies the peptides behind the MS/MS spectra of a run. For each MS2 scan it first determines the charge of the precursor ion from the isotope pattern in the preceding MS1 scan (charge state determination). With the charge, the precursor m/z becomes a peptide mass, and the tool fetches every peptide from the PeptideDB database whose mass lies within LookUpPPM of it. For each candidate it computes the theoretical fragment ions (fragmentation) and compares them with the measured spectrum. Each candidate gets a SEQUEST-like score and an Andromeda-like score. An X!Tandem-like score is computed alongside the Andromeda one. The candidates include reversed decoy peptides, which PSMStatistics later uses to estimate the false discovery rate (what the decoys are for).

Inputs and outputs

All four flags are mandatory. The reader opens files by extension and knows .mzlite and .mzML only, so an mzML file has to carry the extension in exactly that spelling. The tool stops when the database file does not exist.

The tool writes one <run>.psm per input into the output directory. It is a tab separated table with a header. Each row is one candidate peptide for one spectrum with its charge, precursor m/z, theoretical mass, the three scores with their delta values to the next candidates, the peptide sequence and a Label of 1 for a target peptide and -1 for a decoy. The file is opened in append mode, so running the tool twice into the same directory adds a second header and a second set of rows. Delete the old file first. The .psm is read by PSMStatistics. The output directory also receives PeptideSpectrumMatching_log.txt and one <run>_log.txt per input.

Parameters

ChargeStateDeterminationParams is a BioFSharp.Mz ChargeState.ChargeDetermParams record.

Andromeda is a ProteomIQon.Domain.AndromedaParams record.

The default file is peptideSpectrumMatchingParams.json.

Writing a parameter file

open BioFSharp.Mz
open ProteomIQon

let chargeDetermParams : ChargeState.ChargeDetermParams =
    {
        ExpectedMinimalCharge = 2
        ExpectedMaximumCharge = 5
        Width                 = 1.1
        MinIntensity          = 0.15
        DeltaMinIntensity     = 0.3
        NrOfRndSpectra        = 10000
    }

let andromedaParams : Domain.AndromedaParams =
    {
        PMinPMax                = 4, 10
        MatchingIonTolerancePPM = 100.0
    }

let peptideSpectrumMatchingParams : Dto.PeptideSpectrumMatchingParams =
    {
        ChargeStateDeterminationParams = chargeDetermParams
        LookUpPPM                      = 30.0
        nTerminalSeries                = NTerminalSeries.B
        cTerminalSeries                = CTerminalSeries.Y
        Andromeda                      = andromedaParams
    }

// Replace the temp folder with your project folder.
let outputPath = System.IO.Path.Combine(System.IO.Path.GetTempPath(), "peptideSpectrumMatchingParams.json")

Json.serializeAndWrite outputPath peptideSpectrumMatchingParams

Running the tool

Install the tool with dotnet tool install --global ProteomIQon.PeptideSpectrumMatching, then score one run:

proteomiqon-peptidespectrummatching -i path/to/run.mzlite -d path/to/AraTest.db -o path/to/output -p path/to/peptideSpectrumMatchingParams.json

Several runs, three of them scored at the same time:

proteomiqon-peptidespectrummatching -i path/to/run1.mzlite path/to/run2.mzlite path/to/run3.mzlite -d path/to/AraTest.db -o path/to/output -p path/to/peptideSpectrumMatchingParams.json -c 3

All flags:

proteomiqon-peptidespectrummatching --help